BCHM: 4720 Final Exam (cumulative)

Q cycle (Part of complex III function) Cont.

-Bob, Mary, and Sue the story of the 3 Q "Q pool" -Freely mobile in the membrane

How does CoA-SH resemble the phosphopantetheine prosthetic group in ACP?

-Both are covalently attached -Both prosthetic groups are the same -Allows for carrying (free transport) of active groups

Explain the toxic effects of cyanide and carbon monoxide.

-Both inhibit the ability of complex IV to become oxidized. -This decreases flux through the ETC -When flux through the ETC is inhibited proton pumping is disabled and ATP cannot be produced -ATP can't be produced because complex IV is inhibited

Hormonal Regulation of Glycogen Phosphorylase by a Kinase Cascade

-Brain --> adrenal --> epinephrine --> muscle --> binds to epinephrine receptor -Pancreas --> a (fasting) or b (fed) cells --> glucagon or insulin (respective) --> respective receptor -insulin promotes cAMP hydrolysis (cAMP --> AMP) -phosphorylase kinase is activated by Ca2+ (calmodulin) -IP3 (inositol 1,4,5-triphosphate), Ca2+, cAMP/cGMP, and DAG (diacylglycerol) are secondary messenger made in response to hormones -

How do C4 plants work? What are some examples of C4 plants?

-C4 plants work by transporting oxygen to a 2nd compartment to concentrate it so that rubisco doesn't bind oxygen at high temperatures. -CO2 reacts with pyruvates to form a 4 carbon molecule that can be decarboxylated to form Co2 -Some examples are: -tropical plants -grasses -sedges (grassy bush things... think crab grass) -corn

In first 3 Steps of of Calvin Cycle

-CO2 + 4e- --> C1 aldehyde of GAP -2NADPH --> 2NADP+ + 4e- + 2H+ -Oxidation of 2 NADPH drives 4e- reduction of CO2 -Reductive Pentose Phosphate Pathway -NADPH --> NADP

how does calcium binding to calmodulin change its properties? how does calcium binding to calmodulin relieve autoinhibition of a kinase? Is this a catalytic process?

-Ca2+ binds to calmodulin changing the exposure/alignment of the hydrophobic residue -This moves opens up the catalytic site which can then bind to target proteins -calmodulin blocks the catalytic site and auto-inhibits the kinase -Ca2+ then binds and opens up the catalytic site and relieves its auto-inhibition -This process is not catalytic since the kinase has no activity before Ca2+ binds

Question in image on the right:

-Calculating the standard free energy change of a reaction. Answer Below:

Dark reactions:

-Calvin Cycle -Light independent reactions -CO2 -NADPH-> glyceraldehyde 3-P --> glucose --> starch

transport of fatty acid through inner mitochondrial membrane via carnitine (and energy balance)

-Carnitine binds to fatty acyl CoA (favorable) -A translocase (antiporter) can then transport acyl carnitine through the IMM -C8 and shorter can get through the IMM without a transporter

Kinases

-Catalyzes phosphoryl transfer from ATP to a nucleophile acceptor (usually -OH, otherwise -SH/-NH) -Signaling... ~700 kinases in human genome -Random Bi Bi Mech. (seen in picture)

Biosynthetic roles of Citric Acid Cycle

-Cataplerotic Reactions: CAC reaction intermediates depleted for anabolic synthesis (red) -Anaplerotic Reactions: CAC reaction intermediates generated from other metabolites (green) -Shown in the image to the right

how can certain cells 'ignore' hormonal regulation, while others are very responsive to it?

-Certain cells do not have hormonal receptors -(e.g. no insulin/glucagon receptors) -i.e. liver/brain cells have no insulin receptors -Other cells have a high number of these receptors and are thus very responsive -i.e. liver/kidneys have a high number of glucagon receptors

Vectorial Transport in Cytochrome C Oxidase Cont.

-Channels in subunit 1 of Cyt. C oxidase -K channel (Lys 319): mutants block activity completely -D channel (Asp 91): mutants block H+ transport completely... catalysis incompletely -How does e- transfer shift conformers directionality? -Complex IV 2 activities: -Oxidase: 1/2O2 --> H2O -Proton pump

Pigments in Photosynthesis

-Chlorophyll a,b: absorb red and blue light (are green) -Use Mg -carotenoids and phycobilins: absorb blue, green light (are red)

Chloroplast vs. Mitochondria

-Chloroplast: -Weird membrane -Super fluid -galacto lipids -Lots of protein -granae: disks 5 um long -Highly impermeable inner membrane -Outer membrane is permeable -Thylakoid membrane contains the components required for the light reactions of photosynthesis (i.e.): -Light harvesting proteins -Reaction centers -Electron transport centers -ATP synthase -[H+ are pumped into the Thylakoid (Tk) space from stroma

odd chain fatty acid. what is the end product and what do we do with it? (no mechanism)

-Cis d^3-enoyl CoA which cannot be processed by acyl CoA dehydrogenase -Cis d^3-enoyl isomerase converts the double bond into trans d^2-enoyl CoA which is a normal B oxidation substrate

(II) Citrate Synthase

-Citrate (Product inhibition) -Succinyl CoA (Downstream product inhibition) -NADH (Allosteric inhibitor/OA substrate availability

TCA transport proteins:

-Citrate: TCA transport system -Malate/alphaketoglutarate: Malate/alphaKG carrier (antiporter)

Succinyl CoA Synthetase: Cont. Cont.

-CoASH leaves in the formation of succinylphosphate -Succinate leaves active site after histidine binds to succinylphosphate intermediate

Complex III: Coenzyme Q-cytochrome C Oxidoreductase (Cytochrome bc1 complex)

-CoQH2 (ubiquinol) <--> CoQ (ubiquinone) +2e- + 2H+ (-0.045 V) -2 cytochrome c (Fe3+) + 2e- <--> 2 cytochrome c (Fe2+) (0.235 V) - dEo' = 0.0190 V - dGo' = -36.7 kJ/mol (can make ATP)

Two redox cofactors of ETC

-Coenzyme Q: Ubiquinone + 2e- <--> Ubiquinol -Moves in membrane -Cytochrome C: Fe3+ + e- <--> Fe2+ -"skates" on top of membrane -Heme protein -Most conserved protein on earth (very important/ functional) -Small molecules -Moveable

In your own words, explain how some of Dr. Mitchell's personality traits (as portrayed in the 'oxphos wars') made the discovery possible, or how it might have made it more difficult (only use space below).

-Coming from outside of the field without preformed knowledge -unorthodoxy -Confirmation bias that later on prevented hime from accepting other people's views

Complex I: NADH: ubiquinone (CoQ) oxidoreductase Cont. x2

-Complex I generates new structures

Cytochrome B6F Complex from M. Laminosus

-Complex III in ETC -Plastocyonin in right of diagram -Cyt. bf transfers electrons from plastiquinol (QH2) to plastocyanin (Pc) -Protons from plastiquinol are released into the Tk lumen. -Cyt. bf pumps two more protons from the stroma into the lumen (generates a protonmotive force) -Mechanism is similar to the Q cycle of complex III of ETC in cellular respiration -QH2 + 2Pc(Cu2+) --> Q + 2Pc(Cu+) + H+(Tk lumen) -Cu1+ <--> Cu2+ + 1e-

Connection between Glycolysis, TCA cycle, Electron Transport Chain, and Oxidative Phosphorylation

-Complex IV is irreversible -Number of reduced Cyt. C depends on [NADH] -[NADH] in turn regenerates the TCA cycle

Four Complexes of ETC

-Complexes I-IV: I: NADH-Q oxidoreductase II: Succinate-Q reductase III: Q-cytochrome c oxidoreductase IV: Cytochrome C oxidase -Membrane bound big protein complexes -Physically separate assemeblies -diffuse throughout the IMM -Contain multiple components that are not easily separated -Complexes pump protons

Connection between Glycolysis, TCA cycle, Electron Transport Chain, and Oxidative Phosphorylation (summary)

-Connection between the reactions are shown in the diagram on the right -1 NADH --> 10 H+ translocated -3 ATP/H+ should make 3.7 ATP/H+ -Humans have 8 Subunits in the C ring -We make 2.5 ATP/NADH

Strategies used to maintain homeostasis:

-Controlling the amounts of enzymes: -PTM (post translational modification... fast)/allostery -Controlling the accessibility of substrates

Answer:

-Conversion of glycolytic intermediates -Most of the glucose 6 phosphate enters the glycolytic pathway -the oxidative pentose phosphate pathway -All since the cell has much more ribose 5 phosphate than NADPH -Has a low concentration of NADP+ -These conditions may occur in rapidly dividing cells

energy balance of the Cori cycle

-Cori cycle consumes 6 ATP and produces 2 ATP per molecule of glucose/lactate -That means it is energy balance negative

Understand the importance of coupling reactions in the UDP-glucose pyrophosphorylase mechanism and what drives this reaction Cont.

-Coupling the two reactions makes the group transfer favorable

Prosthetic Group

-Covalently bound cofactor

hemiacetal/hemiketal/cyclic hemiacetal

-Created by an aldehyde/ketone reacting with one equivalent of alcohol -Same mechanism as a hydration of a aldehyde or ketone -Endpoint in basic conditions -Can further go onto a ketal or acetal if more equivalents of alcohol were added -nucleophile is alcohol oxygen -electrophile is the carbonyl carbon -cyclic hemiacetal: intramolecular -OH reacts with the carbonyl group

Complex IV: Cytochrome c oxidase Cont. x2

-CuB: -1 copper distorted square planar -proximal to heme a3 -peroxide between heme a3 and CuB

Complex IV: Cytochrome c oxidase Cont. x4

-Cyt C on top of the diagram -O2 reduction to H2O -partial products are dangerous (radicals) -Need 4 e- at once -Goes first to CuA because it's more exposed

Electron Chain in Photosynthetic Bacterial Reaction Center

-Cyt. C(red) starts bound (heme) -QA (quinone) is bound after step 3 -After step 5 the e- comes from QB from Cyt. bc -Cyt. bc1 is like complex III in the ETC -No oxygen produced due to cyclic e- transport -Cyt. C: water soluble -Anaerobe: an-oxygenic (no O2 production) -End Game: [H+] gradient to power F1F0 ATPase -The possible return of e- from pheophytin to the oxidized special pair is suppressed by the "hole" in the special pair being refilled by an e- from the cytochrome subunit and the e- from the pheophytin being transferred to quinone (QA) that is farther away from the special pair (steps 3 and 4). -The reduction of quinone QB on the cytoplasmic side of the membrane results in the uptake of two protons from the cytoplasm (steps 5 and 6)

Complex IV: Cytochrome c oxidase Cont. x3

-Cyt. c oxidase consists of 13 subunits -4 Cyt. C reduced -cytochrome c --> CuA/CuA --> heme a --> heme a3 --> CuB

What is the human equivalent of plastocyanin?

-Cytochrome C is the human equivalent of plastocyanin because it works to shuttle electrons (like plastocyanin)

Plant Photosystem (Complex II)

-Cytochrome b6-F complex: Cyt. b6, Cyt. F, Fe-S proteins -Mediated by plastoquinone (Q) (quinone analog)

Aldolase Cont.

-DHAP: dihydroxyacetone phosphate -GAP: Glyceraldehyde 3-phosphate (former C4-C6 -Enolate: resonance stabilized intermediate -Aldol cleavage before C3-C4 requires carbonyl at C2 and hydroxyl at C4 -DHAP further broken down into GAP by triosphosphate isomerase

Final 3 reactions (8-10 Overall)

-Decarboxylation of pyruvate -decarboxylation needs a ketone at Ca or Cb (better) -With C3 we can only put a group on Ca -Phosphate needs to be made a good leaving group in acetyl-CoA

Notable non-redox reactions:

-Dehydration/rehydration -Protonation/deprotonation

Esterification Reactions are Catalyzed by Various Acyl Glycerol Transferase

-Diesterification is catalyzed by various lipases: -Monoglyceride lipase -diglyceride lipase -Triglyceride lipase -Phospholipase -Hormonally controlled with some specificity -Olestra: A fat substitute that cannot be digested -Sucrose octo-ester

Dietary Fats Ingested as Triglycerides

-Dietary fats are emulsified in the small intestine by bile salts -e.g. taurocholic acid -Hydrolysis of triglycerides to fatty acids, mono and diglycerides -These can then be absorbed into the intestinal mucosa (epithelial cells lining the intestine)

Dimer of Mitochondrial ATP Synthase

-Dimer of 2 F1F0 ATPase complex -C E Y: rotor rotates within a b hexamer held by a b2 d

Glycogen Phosphorylase X-Ray Structure

-Dimer: monomers related by 2 fold symmetry -Catalytic site: G1P bound pyridoxal phosphate (P1P) -Pyridoxal S' phosphate: PLP, vitamin B6 -PLP phosphorylase: Schiff's base -Glycogen binding site: Maltoheptose bound -30 A from catalytic site -Allosteric effector site: -AMP: activator -ATP, G6P: inhibitors -Nucleotide inhibitor site: -adenosine, caffeine: inhibitors -Phosphorylation site: Ser14

DNP

-Dinitrophenol -Active ingredient in some herbicides and pesticides -Used as a weight loss drug -Banned in 1938 by the FDA -Comeback in popularity online and sales soared in 2014 -Causes extreme overheating -People died and in 2019 people went to jail. -

(4) Branching Enzyme (amylo(1,4-->1,6)transglycosylase)

-Distinct from debranching enzyme -Cleaves a(1-->4) -Makes a(1-->6) -Length of transferred segments is roughly 7 residues -Creates new non-reducing ends -Branch points every 8-12 residues -This enzyme can count and measure

Potential unsolved problem candidate

-Do both substrates need to be bound to hexokinase to induce conformational change? -By understanding whether both substrates need to be bound to induce conformational change one can make a more accurate induced fit model. Making a more accurate model for hexokinase would allow researchers to better understand the specificity of hexokinase and likely other kinases.

C Ring rotation

-Does the C ring move unidirectionally? -Unidirectional in the CCW direction. -C ring rotates because the H+ gradient provides continuous protonation -Full rotation produces 3 ATP -Number of subunits in the C ring determines the efficiency of the machine -Human: 8 subunits -Bacteria: 13 subunits -Motors are faster with more subunits -The fewer subunits the more efficient the motor efficiency = 1/N (N = # of subunits). Each 360 degree turn produces 3 ATP so fewer subunits means less H+ needed to produce each H+

Noncyclic Electron Transport

-Driven by hv --> PSII, PSI -4e- released from H2O --> O2 -2NADP+ reduced to 2NADPH -Photophosphorylation drives ATP synthesis through establishment of H+ gradient

(I) Pyruvate Dehydrogenase Complex

-E2: Dihydrolipoyl Transacetylase -Acetyl CoA (Product inhibition) -E3: Dihydrolipoyl Dehydrogenase -NADH (Product inhibition) -Both Reactions reversible

What does the proton gradient drive?

-Electron potential (dE) -Heat production -Active transport -Flagellar rotation -ATP (~P) -NADPH Synthesis (plants)

Photophosphorylation

-Electron transport drives H+ transport -stroma --> lumen -[H+]L/[H+]S >> 1 -Plastiquinone (Qox)--> plastisemiquinone (QH.)--> plastoquinol (Hydroquinone) (QH2) -Q Cycle

Fun ETC Facts:

-Electrons do not flow through the ETC unless there is ADP available to be converted to ATP -Respiratory (acceptor) control: regulation of Oxidative Phosphorylation by ADP. -Inhibitory Factor 1 (IF1): prevents ATP hydrolysis when oxygen is not available to accept the electron of the ETC. -Non-shivering thermogenesis: -UCP-1 (conserved) dissipates proton gradient (fatty acids) in brown adipose tissue (rich in mitochondria). -Uncouples oxidative phosphorylation and results in heat production instead of ATP -Brown adipose tissue is present in hibernators (e.g. marmots/bears)

Electron Transport Chain (ETC)

-Electrons passed through protein/cofactor complexes -Complexes convert NADH or FADH2 oxidation to a gradient of H+ -[H+] IMS >> [H+] matrix -Energy from the proton gradient is used to generate ATP -This coupling is called: oxidative phosphorylation -Oxidation of 1 NADH ~= 2.5 ATP -Oxidation of 1 FADH2 ~= 1.5 ATP -Why is there slippage (i.e. nonstoichiometry) in this process? - ~30% efficient -Under physical conditions ~70% efficient

know that only C16 or shorter saturated FA are made by FAS, longer and unsaturated chains are made by elongases and desaturases, respectively Cont.

-Elognases use malonyl CoA or acetyl CoA -Cn --> Cn+2

Triacylglycerols, with their hydrocarbon-like fatty acids, have the highest energy content of the major nutrients. Answer the questions about energy reserves in adipose tissue. Assuming that 15% of the body mass of a 70.0 kg adult consists of triacylglycerols, calculate the total energy reserve, in units of kilojoules and kilocalories, that is available from triacylglycerols. If the basal energy requirement is approximately 8,400 kJ/day (2,000 kcal/day), calculate the amount of time that this 70.0 kg person could survive if the oxidation of fatty acids stored as triacylglycerols were this person's only source of energy. Calculate the weight loss in pounds per day under the starvation conditions in the previous part (1 lb=0.454kg)(1 lb=0.454kg) .

-Energy reserve: 398960 kJ = 95445 kcal -Time until reserves are gone: 47.72 -Weight loss per day: 0.485 lbs/day

TCA Cycle Facts:

-Entry point for AA and porphyrin degradation -Building blocks for AA and porphyrins -Entry point for FA Degradation -Almost all C from CHO Degradation --> Acetyl-CoA --> C as CO2 eliminated -CO2 released from oxaloacetate and not acetyl CoA -Carbons in acetyl CoA regenerate new oxaloacetate -No net synthesis of OAA (oaxaloacetate) in TCA cycle

TCA cycle: Krebs Cycle/Citric acid cycle (Mechanisms):

-Entry point for: FA, CHO, and AA catabolism -Metabolic precursors for AAs, nucleotides, cofactors, neurotransmitters, etc. -Pyruvic acid combines with coenzyme A to form Acetyl-CoA which enters Krebs cycle -in matrix of mitochondria: transported to mitochondria through pyruvate [H+] transporter - each turn of Krebs cycle produces 1 molecule of ATP, 1 molecule of FADH2, and 3 molecules of NADH -byproduct CO2 is exhaled oxidized: citrate reduced: NAD+ and FAD+

Phosphoglycerate mutase Cont. Cont.

-Enzyme needs to be reprimed with 2,3 BPG in step 3 -Need to remove the regulator (phosphatase) since wasting 1 ATP to make the regulator.

all life form use glucose polymers for energy storage

-Eukaryotes, archea, and bacteria all use glycogen as a glucose polymer for energy storage -Plants uses starch as a glucose polymer for energy storage

Diagram of Fat Metabolism Through Various Organs

(1) Bile salts emulsify dietary fats in the small intestine making micelles (2) Intestinal lipases degrade triacylglycerols -(water-lipid interface) (3) Fatty acids and other breakdown products are taken up by the intestinal mucosa and converted into triacylglycerols (4) Triacylglycerols are incorporated with cholesterol and apolipoproteins into chylomicrons (5) Chylomicrons move through the lymphatic system and bloodstream to tissues (6) Lipoprotein lipase activated by apoC-II in the capillary converts triacylglycerols to fatty acids and glycerol (7) Fatty acids enter cells through diffusion (8) Fatty acids are oxidized as fuel or reesterified for storage -Myocyte: storage for muscle cells -Adipocyte: storage for fat cells

Sources of Fat

(1) Dietary Fats: -fatty acids --> acetyl CoA or triglycerides -fatty acids --> muscle --> fatty acids --> Acetyl CoA (through b oxidation) --> CO2, H2O, and ATP (through TCA and Oxidative phosphorylation) -fatty acids --> adipose (fat cells) --> fatty acids --> triglycerides (storage) (2) Stored Fats: -glycerol --> glycolysis -Triglycerides --> fatty acids (through lipases) --> Blood (hormonal control here) --> skeletal and cardiac muscle (3) De Novo Synthesis (liver): - carbohydrates + amino acids --> acetyl CoA - --> ketone bodies (source of energy) --> blood - --> fatty acids --> triglycerides --> lipoproteins --> blood -Everything is more clear in the image

Fate of pyruvate / TCA cycle at high acetyl CoA, NADH, ATP; at low acetyl CoA, NADH, ATP; and at high acetyl CoA, low NADH and low ATP.

(1) High: Acetyl CoA, NADH, and ATP -Lots of energy -pyruvate --> glucose is enhanced and excess glucose is made into glycogen (storage) -TCA down -Pyruvate decarboxylase up (acetyl CoA) (2) Low: Acetyl CoA, NADH, and ATP -low energy -pyruvate --> glucose is suppressed -Pyruvate carboxylase down -TCA up (PDH) (3) High: Acetyl CoA Low: NADH and ATP -pyruvate carboxylase (PC) up (acetyl CoA) -acetyl CoA --> OAA --> TCA cycle -TCA up -No glucogenesis

Glycolysis: overall reaction

(Glucose) + (2 NAD+) + (2ADP) + (2Pi) --> (2 pyruvate) + (2NADH) + (2ATP) + (2H2O)

Preventing hydrolysis of ATP

(likely answers): -Through pH (physiological pH allow ATP to be stable) (higher pH = less hydrolysis) -Limiting the presence of Mg2+ ions (needed in hydrolysis)

Matrix

- <50% water (very gloopy) -contains enzymes of the CAC and pyruvate dehydrogenase complex

Percentage of Oxygen generated and ATP produced

- >80% of oxygen used - >80% of ATP produced

Write out the net reaction of the PPP if the cell needs NADPH and Ribose-5-Phosphate.

- Glucose 6 phosphate + 2 NADP+ + H2O --> ribose 5 phosphate + 2NADPH + 2H+ + 2CO2

GPCR: what are they, how do they work. Remember that there are MANY different GPCR, activated by a wide variety of hormones

- Heterotrimeric G-protein coupled receptor -Substrate binds to the G protein receptor and GTP is released -GTP to coupled enzyme and it is activated -G-protein hydrolyzes GTP to GDP and goes back to inactive state

Describe how the pentose phosphate pathway (PPP) can generate glycolytic intermediates.

- PPP can synthesize glycolytic intermediates through stage 1 (generating Ru5P) -Ru5P will then undergo stage 2 (R5P to E4P) -These will both undergo carbon shuffling reactions in stage 3 -Will ultimately form glycolytic intermediates such as GAP or F6P

Proton Motive Force (PMF)term-56

- dG(ims --> mat) = -dG(mat --> ims) - dP is the measure of dG(ims --> mat) in electrochemical units -membrane thickness of 80 A -lots of equations in the image -Typically: -psi(mat) - psi(ims) = -168 mV -pH(mat) -pH(ims) = 0.75 - dG(ims --> mat) = -20.4 kJ/mol per proton -Need more than 1 H+ to produce 1 ATP (need 3H+/ATP)

The difference in pH between the internal and external side of the mitochondrial membrane is 1.4 pH units (the external side is more acidic). If the membrane potential is 0.06 V (inside negative), what is the free energy released on transporting 1 mol of protons back across the membrane (from out to in). Assume a temperature of 310 K.

- dG(in --> out) = F(psi(out) - psi(in)) - 2.3RT(pH(out) -pH(in)) -psi: membrane potential

ATP produced from complexes

- dGo' = I + III + IV = -218 kJ/mol ~ 2.5 ATP - dGo' = II + III + IV = -148 kJ/mol ~ 215 ATP

F1F0 ATP Synthase (Complex IV)

-"Lollipop" structures inner mitochondrial membrane protruding into the matrix -F0: -water insoluble transmembrane complex -Proton translocation channel -F1: -Water soluble complex -ATP synthesis (ADP + Pi <--> ATP + H2O) -Middle and peripheral stalks (anchor point)

Calvin Cycle and PPP

-"mirror image" -Many enzymes common to the calvin cycle and the non-oxidative stage of PPP -More similarities in the image

Regulation by enzyme covalent modification

-"post translational" modification -Protein phosphorylation: -catalyzed by protein kinases (Highly specific) -Removed by protein phosphatase (unspecific)

how are ketone bodies converted into energy? How does liver avoid consuming ketone bodies

-"selfless liver" -CoA transferase is not in the liver so the liver cannot consume ketone bodies -Can only produce ketone bodies -Liver cannot convert ketone bodies back into acetyl CoA -Liver does not have OAA so it cannot process acetyl CoA

Phosphorylase Kinase Structure

-(alpha, beta, gamma, delta)4 composition -active site on gamma subunit -Phosphorylase kinase is activated first by Ca2+ binding and then by phosphorylation (by PKA) -delta subunit (calmodulin) is the calcium sensor -Phosphorylase kinase is maximally effective when phosphorylated and Ca2+ is bound

Glucose-alanine cycle to transport more carbon to the liver for gluconeogenesis

-1 Glucose is made into 2 pyruvate via glycolysis -pyruvate cannot be transported to it is made into alanine (can be transported) -2 alanine is then made back into 2 pyruvate in by alanine aminotransferase -2 Pyruvate is made into 1 glucose by guconeogenesis -alanine (at first pyruvate) transports more carbon to the liver

b. How many moles of ATP could it theoretically synthesize per mole of NADH, assuming the reaction is 100 % efficient?

-1 NADH supports the transport of roughly 10 H+ so only 2 ATP can be made -Less efficient than humans -Humans in theory only use the energy from the translocation of 3 H+ per ATP -Humans make ~ 3.3 ATP per NADH

Overall B Oxidation of Palmitoyl CoA

-10.5 ATP formed from the 7 FADH2 (1.5*n FADH2) -17.5 ATP from the 7 NADH (2.5*n NADH) -80 from the 8 acetyl CoA molecules (10*n) (TCA cycle) -2 molecules are consumed and split into AMP and 2 molecules of orthophosphate -Complete oxidation yields 106 ATP

Cross Linked Polymer

-12 layers/fractals - ~60,000 glucose units/particle -Branch points separated by 8-12 glucose units -One reducing end -Many nonreducing ends (indicated as N in the image) -Stored in cytoplasm as glycogen granules Modeling: -Max # of glucose/volume -Max # of ends -Minimal repulsion

Cori Cycle

-1947 Nobel prize in medecine -Gerty was the first American Woman to win a nobel prize in science -Lactate and alanine formed by contracting muscles are used by other organs -Muscle and liver display interorgan cooperation in a series of reactions called the Cori cycle -Liver takes up lactate and converts it into glucose --> blood -Pyruvate accepts NH2 from amino acids --> alanine -muscle wasting disease -Alternately in the liver: alanine --> pyruvate -Alanine aminotransferase ALT (diagnostic) -abundant liver enzyme

Glycolysis Overall Reaction Comparison

-2 P gained -If both glycolysis and gluconeogenesis are active at the same time then its a futile reaction (no real gain) -Both must not be active at the same time

Complex IV: Cytochrome c oxidase (Cox)

-2 cytochrome c (Fe2+) <--> 2 cytochrome c (Fe3+) + 2e- (-0.235 V) -1/2O2 + 2e- + 2H+ <--> H2O (0.815 V) - dEo' = 0.580 V - dGo' = -112 kJ/mol (can make ATP) -ATP is not directly made from this [H+] gradient

Transport of PEP and OAA from Mitochondrion to Cytosol

-2 routes for OAA "swing king" -OAA must be first converted to aspartate through aspartate aminotransferase or to malate by malate dehydrogenase.

Answer

-200.7

Gluconeogenesis Pathway Cont.

-2nd half (after aldolase) is at equilibrium -lactate comes from muscle -dG in direction of gluconeogenesis -Bypasses 3 reactions --> reciprocal regulation

how is cAMP-dependent protein kinase activated by cAMP?

-4 cAMP bind to the 2 regulatory subunits which then change conformation releasing 2 catalytic cAMP dependent protein kinase residues

Percentage of glucose made through gluconeogenesis

-50% of the glucose made in the last 20 hours is made through gluconeogenesis

E3 Dihydrolipoyl dehydrogenase (part 1)

-6 Steps in this reaction -Dihydrolipoamide is oxidized by way of NADH to form lipoamide -E3 has two adjacent Cys-SH

Efficiency of Light Reactions and Overall Reaction

-8 photons = 8 * 170 kJ/mol = 1300 kJ/mol (input) -560 kJ/mol for the overall reaction -8 photons are required to yield 2 molecules of NADPH and 3 molecules of ATP -In cyclic phophorylation: two photons yield one molecule of ATP and no NADPH

A fatty acid composed of 18 carbon atoms undergoes b oxidation. How many acetyl CoA, FADH2}, and NADH does b oxidation of this fatty acid generate? Calculate the net ATP generated by the B oxidation of the 18 carbon fatty acid. Assume that each FADH2 generates 1.5 ATP and each NADH generates

-9 acetyl CoA, 8 FADH2, 8 NADH -120 ATP

(b) How efficient is this process compared to the energy of an einstein (mole of photons) at a wavelength 680nm? Use E=​hc/λ.

-98.6% efficient -Measured / theoretical

Concept of Substrate Cycles

-A pair of reactions such as phosphorylation of fructose 6 phosphate to fructose 1,6 biphosphate and back is called a substrate cycle. -Both reactions are not fully active at the same time because of reciprocal allosteric controls -There can be some detectable activity of opposing pathways futile cycle -Substrate cycles are biologically important -Enhance metabolic signals -A small change in two opposing cycles can lead to a large change in the net flux -20% change in activation on both sides leads to a 380% change in flux.

What is the ETC and oxidative phosphorylation?

-A series of oxidation reduction reactions using NADH and FADH2 as the initial electrons donors and molecular oxygen as the terminal electron acceptor -Initial electron donors: NADH and FADH2 -Terminal electron acceptor: Molecular oxygen -Movement of electrons through complexes I through IV will generate a proton gradient -->

What is a second messenger - and name at least two

-A substrate made in response to hormones -cAMP -cGMP -DAG -IP3

two forms of ACC - regulatory role of ACC2 in making malonyl CoA to prevent the import of FA into mitochondria

-ACC1: In adipose -makes fatty acid -ACC2: most tissues -makes malonyl CoA for regulatory reasons -Inhibits fatty acid import into the mitochondria

activation of acetyl coA by ACC - role of biotin, why carboxylation, mechanism. what is the fate of the HCO3 that is being put on?

-ACC2: In most tissues it inhibits fatty acid transport into the mitochondria -ACC1: in adipose cells and makes fatty acids -biotin serves as the carboxy phosphorylate intermediate -allows for the transfer of HCO3- to acetyl CoA -HCO3- ends up on acetyl CoA and makes it into malonyl CoA

Plant Photosystem (Complex V)

-ATP Synthase: -H+ gradient drives ADP + Pi --> ATP -Photophosphorylation -[H+]lumen >> [H+]stroma

Photophosphorylation

-ATP synthesis driven by the protonmotive force -Very similar to animals

Nucleotide transport proteins

-ATP, ADP: Adenine nucleotide translocase (antiporter) -H2PO4-, H+: Phosphate translocase (symporter)

Activated carriers:

-ATP: activated carrier for phosphoryl group -NAD+/NADH, NADP+/NADPH, FAD/FADH2: activated carriers of electrons -Coenzyme A: activated carrier of 2 carbon fragments

Substrates that do not make glucose in gluconeogenesis

-Acetyl CoA -Leucine -Lysine

No Defined transport

-Acetyl CoA -NAD+/NADH -Oxaloacetate -"smuggle" them across

Fat Burns in the Flame of Carbohydrates

-Acetyl CoA + OAA --> citrate (TCA cycle) -If no OAA then OAA --> gluconeogenesis -Acetyl CoA is diverted into ketones -transport for acetyl CoA

Acetyl CoA in the Citric acid Cycle

-Acetyl CoA --> CoA-SH + CO2 -Coupled to the synthesis of 3 NADH, 1 FADH, 1 GTP

Reciprocal Regulation of Gluconeogenesis

-Acetyl CoA is a feedback inhibitor of pyruvate dehydrogenase (PDH) -NADH and ATP suppress the TCA cycle -Isocitrate and aketoglutarase repression -produced inhibition (1) High: Acetyl CoA, NADH, and ATP -Lots of energy -pyruvate --> glucose is enhanced and excess glucose is made into glycogen (storage) -TCA down -Pyruvate decarboxylase up (acetyl CoA) (2) Low: Acetyl CoA, NADH, and ATP -low energy -pyruvate --> glucose is suppressed -Pyruvate carboxylase down -TCA up (PDH) (3) High: Acetyl CoA Low: NADH and ATP -pyruvate carboxylase (PC) up (acetyl CoA) -acetyl CoA --> OAA --> TCA cycle -TCA up -No glucogenesis

Diagram of Pyruvate Carboxylase Subunit Cont.

-Acetyl CoA is an allosteric regulator (does not participate in the reaction) -Activated CO2 (carbon) is on biotin --> [flexible arm] swings into CT domain to donate pyruvate (C3) to OAA (C4)

E2: Dihydrolipoyl transacetylase Cont.

-Acetyl CoA is formed in the breakage of the thioester bond -Dihydrolipoamide is reduced and for further reactions it needs to be oxidized

Yeast Pyruvate Kinase

-Activated by F 1,6 BP -Inhibited by ATP -Tetramer: active -Dimer: inactive

What activates Rubisco? How does this link the dark reactions to the light reactions in photosynthesis?

-Activated by: -Mg2+ -thioredoxin -modified lysine carbamate (which binds Mg2+) -pH of 8 -ATP -These link rubsico to the light reactions since as photosynthesis continues the pH and Mg2+ concentration in the stroma increases.

Covalent modification of Enzymes

-Activation -Inhibition

Methods of transport in the Mitochondria

-Active transport (shown in image) -Facilitated diffusion -Simple diffusion

Various Phosphofructokinase (PFK1) proteins Cont.

-Active: R162 points at F6P allowing it to bind -repressed?: E161 prevents substrate binding -ADP promotes R state (relaxed)

TCA Cycle: CO2 from the first pass

-After one pass CO2 comes from Oxaloacetate (NOT Acetyl CoA)

Stage 1 of TCA cycle Diagram

-Again pyruvate --> acetyl-CoA -Bottom right is the pyruvate dehydrogenase complex of E. Coli: -E1 in yellow -E2 in green -E3 in red -Table showing functions of the subunits is shown below

Glycogen Breakdown Overview

-All enzymes are bound to granules (1) glycogen(n+1) [a(1-->4)] + Pi --glycogen phosphorylase--> glycogen(n) + glucose 1 phosphate (2) Glucose 1 phosphate <--Phosphoglucomutase--> Glucose 6 phosphate (3) Glucose 6 Phosphate + H2O --glucose 6 phosphatase--> glucose + Pi (liver only) (4) Debranching enzyme [a(1-->6) linkage]

Two Types of Reciprocal Regulation

-Allosteric -Hormonal -Acetyl CoA activates pyruvate carboxylase

Succinyl CoA Synthetase: Nucleotide diphosphate kinase

-Also through phosphohistidine intermediate -GTP + ADP <--> GDP + ATP -remaining 3 steps regenerate oxaloacetate -Unusual since: 1) membrane bound (part of etc) 2) FAD not NAD+ as e- acceptor (Hydride transfer)

Ketone Bodies (Ketogenesis)

-Alternative fuel source when energy reserves are low -In the liver -Fatty acids --> Acetyl CoA - --> ATP (through TCA) - --> acetoacetate or 3-hydroxybutyrate (ketone bodies) --> [Muscle/brain] ketones --> acetyl CoA --> ATP -Transport system for acetyl CoA -Ketoacidosis: Overproduction of ketones -Ketosis: high levels of blood ketones -acidosis: decreased blood pH -acetone production from acetoacetate -Severe starvation: -Gluconeogenesis depletes OAA driving acetyl CoA to OAA -Juvenile Diabetes (type 1): -insufficient glucose uptake -elevated gluconeogensis -Elevated B oxidation -Blood ketones: -90mg/100ml (untreated) -3mg/100ml (normal) -A keto-diet does not cause ketoacidosis

steps of beta oxidation in mitochondria.

-An activated fatty acid is oxidized to introduce a double bond -The double bond is hydrated to introduce a hydroxyl -The alcohol is oxidized to a ketone -The fatty acid is cleaved by coenzyme A to yield acteyl CoA and a fatty acid chain 2 carbons shorter

Ketone bodies: what are they, when and where (which organ) are they generated? what is their purpose and fate?

-An alternative fuel source when energy reserves are low -ketogenesis -Made in the liver -Acetyl CoA is diverted into ketones (transport form of acetyl CoA) which is then transported to the muscle/brain -the ketones are then made into acetyl CoA in muscle or the brain which then is used to make ATP -Fate: ATP production in brain/muscle

What is trans-fat?

-An unsaturated fat, formed artificially during hydrogenation of oils, containing one or more trans double bonds.

Pyruvate Carboxylase Cont.

-Anaplerotic reaction (replenishes TCA intermediates) -Occurs in mitochondria -Activated by Acetyl CoA -TCA cycle is empty (Acetyl CoA has no place to go) -Needs OAA

How animals and plants generally produce energy

-Animals: O2 + CHO --> CO2 + NADH + ATP + H20 -Plants: sunlight (hv) + H2O --> O2 + ATP + NADPH NADPH -CO2-> CHO (starches, sugars, cellulose)

Light Harvesting Antennae

-Antennae: Light harvesting complexes -In Tk membrane photosynthetic reaction centers lie next to antennae

Arsenate poisoning

-Arsenite: an inhibitor of E2 -used to be a miracle cure and treat syphilis in 1800s -Arsenate irreversible binds to dihydrolipoamide -some sulfhydryl reagents (2,3-dimercapto-ethanol) relieve inhibition by forming a complex with arsenite that can be excreted

Steps 6 and 7 (overall)

-At ATP break even point -What is the driving force for converting GAP --> 3PG: -The greater phosphoryl group transfer potential of GAP compared to ATP drives the synthesis of ATP and, thus, the reaction. -ADP --> ATP -Oxidation of GAP energy is trapped as 1,3-BPG

A Special Pair of Chromosomes Initiate Charge Separation (Bacteria)

-Bacterial Photosynthetic reaction center -No Mg2+ -Absorbs infrared -Absorbs at 360 nm -Electron carrying prosthetic groups begin at bacteriochlorophylls and end at a bound quinone

Structure of Phosphofructokinase 2

-Bidirectional enzyme -Toggled by phosphorylation of serine

What are bile salts and what is their purpose

-Bile salts breakdown large globules of fat into smaller droplets of fat -essentially act as bodily detergents -Bile salts also help the body absorb those droplets

Arrange the four major steps in the elongation of fatty acid chains by fatty acid synthase.

-Binding of malonyl‑CoA to ACP -Condensation by B-ketoacyl synthase -Reduction by B-ketoacyl reductase -Dehydration by 3-hydroxyacyl dehydratase -Reduction by enoyl reductase -Butyryl group transfer from ACP to β‑ketoacyl synthase

(1) Glycogen Phosphorylase

-Binds to glycogen + Pi -acid-base catalysis --> cleavage of a(1-->4) link -pyridoxal phosphate (PLP) cofactor -Oxonium ion (oxygen ion with 3 bounds) intermediate that is resonance stabilized. -Group transfer to Pi (makes glucose 1 phosphate) -retention of configuration -We are making a high energy P bond by using the energy from breaking the glycosidic bond

Diagram of Pyruvate Carboxylase Subunit

-Biotin carboxylase is a tetramer with each monomer having 2 active sights -Enzyme is only active in presence of Acetyl CoA -CT: carboxyl transferase -Acetyl CoA binding sight on PT (pyruvate carboxylase tetramerization domain)

(a) In a plant photosystem, P680 in its ground state has ​reduction potential​ +1.0 V. Excitation of P680 by light decreases the reduction potential to -0.8 V. Calculate o'​ΔG​ ​in kJ/mol for this shift in reduction potential. (b) How efficient is this process compared to the energy of an einstein (mole of photons) at a wavelength 680nm? Use E=​hc/λ.

-Excitation greatly facilitates p680 oxidation -Same equation as above

Allosteric Control:

-F 2,6 biphosphate, AMP, citrate --> TCA cycle reactions -Acetyl CoA activates PDH kinase (remember TCA cycle) inactivates PDH

F0 and F1 ATPase structures

-F0 is membrane bound -C ring is in the top right of the diagram -cytoplasm at bottom of the page

F1F0 ATP Synthase (Complex IV) Cont.

-F0: C subunits arranged in "C ring" (~8-15 C subunits) -F1: a3b3 alternating heterohexamer -a subunits: regulatory ATP bound -b subunits: catalytic (empty, ADP, ATP bound) -In the diagram: -C-E-gamma subunit is the rotor and colored yellow -Red subunit is the stator and stalk -The rotation of the gamma subunit inside the a3 b3 hexamer causes the domain movements in the B subunits

What is used as an electron acceptor for going from alkane to alkene (deja vu all over again)

-FAD is used as the electron acceptor from alkane to alkene

Complex II: Succinate dehydrogenase

-FADH2 <--> FAD + 2e- + 2H+ (0.040 V) -CoQ (ubiquinone) + 2e- + 2H+ <--> CoQH2 (ubiquinol) (0.045 V) - dEo' = 0.085 V - dGo' = -16.4 kJ/mol (No ATP) -TCA cycle

know that only C16 or shorter saturated FA are made by FAS, longer and unsaturated chains are made by elongases and desaturases, respectively

-FAS cannot make past C16 -mammals lack enzymes that induce double bonds past C9

Complex I: NADH: ubiquinone (CoQ) oxidoreductase Cont. x3

-FMN: -like FAD - e- and H+ from NADH -Fe-S cluster's e- transfer potential depends on geometry

substrate concentration

-Fast -Affects flux -e.g. : Feed forward

covalent modification

-Fast -Phosphorylation and others -often hormone induced with cascades of signaling

Fat Metabolism Overview

-Fat esterification of OH will produce fatty acids -e.g.: glycerol --> monoacylglycerol --> diacylglycerol --> triacylglycerol -Same thing happens with phospholipids

Complex III: Coenzyme Q-cytochrome C Oxidoreductase (Cytochrome bc1 complex) Cont.

-Fate of ubiquinol generated by complex I and II -QH2 + 2Cyt C(ox) + 2H+(matrix) --> Q + 2Cyt C(red) + 4H+(intermembrane space) -prosthetic groups: -Two Heme b: cytochrome b562 (bh) cytochrome b566 (bl) -One Heme c: cytochrome c1 -One [Fe-S] cluster: Rieske iron sulfur protein ISP -Vectorial transport of 4H+/2e-

What is fat, what are phospholipids?

-Fats are fatty acids used for energy storage -Phospholipids have a hydrophilic head group and form membranes and micelles for transport.

the pathway of dietary fat until it reaches its target tissue

-Fats ingested -Bile salts emulsify dietary fats in the small intestine -Fatty acids are taken up by the intestinal mucosa and converted into triacylglycerols -Triacylglycerols are made into chylomicrons -Lipoprotein lipase converts triacylglycerols to fatty acids and glycerol -Fatty acids can then enter the cell

Place the reactions and relevant locations of reactants for the degradation of an even‑chain saturated fatty acid in the proper order.

-Fatty acid in the cytoplasm -Activation of fatty acid by joining to CoA -Formation of carnitine ester -Acyl CoA in mitochondria matrix -FAD liked oxidation -Hydration by enoyl CoA hydratase -NAD+ linked oxidation -Thiolytic cleavage by B-ketothiolase -Acetyl CoA enters the citric acid cycle

overall similarities of fatty acid degradation (beta oxidation) and fatty acid synthesis

-Fatty acid synthesis and degradation are both four steps and the reverse of each other -Both deal in 2C subunits

Fatty Acid Degradation and Synthesis Mirror Each Other in Their Chemical Reactions

-Fatty acid synthesis and degradation consist of four steps that are the revers of each other. -Fatty acid degradation: -an oxidative process that yields acetyl CoA -an activated fatty acid is oxidized to introduce a double bond -the double bond is hydrated to introduce a hydroxyl group -The alcohol is oxidized to a ketone -The fatty acid is cleaved by coenzyme A to yield acetyl CoA and a fatty acid chain 2 carbons shorter -Fatty acid synthesis: a reductive process that begins with malonyl CoA (a modified version of acetyl CoA) -activated acyl group is fused to activated malonyl group (C3) (coupled to decarboxylation --> CO2 to power the reaction) -Carbonyl group is reduced to a methylene group -Both deal in 2 carbon units

Fatty Acid Oxidation

-Fatty acids --> acetyl CoA -C16 --> 8C2 (1) Activation of fatty acids by addition of CoA -product in the cytosol -High energy activated bond (2) Transport of fatty acyl Coterm-271A to Mitochondria (3) Cleavage of Ca --> Cb bonds: -Products: acetyl CoA, NADH, FADH2 (4) Oxidative Phosphorylation coupled to electron transport -NADH, FADH2 --> ATP

Why are fatty acid more energy rich (per gram and per carbon) than glycogen?

-Fatty acids are more reduced than sugars and thus carry more energy -There are more electrons around each carbon -When electrons move to more electronegative atoms (i.e. C --> O) then energy is released. So the more electrons available the more energy is released

Packaging into Chylomicrons

-Fatty acids are reesterified into triacylglycerides Chylomicrons: -lipid protein aggregates -Outside: apoproteins, phospholipids, cholesterols, and cholesterol esters -Inside: Triglycerides -apoproteins are recognized by cell receptors - 100-500 nm in size -Similar to HDL and LDL -Polar head groups of phospholipids face to the outside -Chylomicrons are transported to the blood -Targeted to certain tissues (especially muscles and fat)

(2) Transport of Fatty Acyl CoA to the Mitochondria

-Fatty acyl CoA + Carnitine --> Fatty acyl carnitine -carnitine palmitoyl transferase I, II is the enzyme involved -This is a Favorable reaction -Carnitine is a zwitterion -Antiporter moves fatty acyl CoA into and out of the inner mitochondrial matrix -C8 and shorter can enter the IMM without a transporter

Metabolite binding inhibition

-Feedback inhibition -Direct product inhibition -Downstream product inhibition via competition, allostery, substrate availability

Ferrodoxin/NADP+ Reductase Facts

-Ferrodoxin and NADP+ reductase transfer electrons from ferrodoxin (2xe-) to form NADPH (2e-) -Formation of NADPH occurs on the stromal side of the Tk membrane --> carbohydrate synthesis -Electron flow from H2O to NADP+ is called the Z scheme of photosynthesis -Pc(Cu2+) + Fd(ox) -hv-> Pc(cu+) + Fd(red) -dG0' = 75 kJ/mol -Need light because its unfavorable -Cyclic: 2H+/e- * 1e-/hv * 1 ATP/3H+ --> 2/3 ATP/hv -Z scheme: 12H+/4e- * 1e-/2hv * 1 ATP/3H+ --> 1/2 ATP/hv -Z scheme is more efficient per photon

in broad steps, how are acetoacetate and hydroxybutyrate made from acetyl CoA?

-Formed from acetyl CoA in the liver (1) 3-ketothiolase (2) hydroxymethylglutaryl CoA synthase (3) hydroxymethylglutaryl CoA cleavage enzyme (4) d-3-hydroxybutyrate dehydrogenase -acetoacetate spontaneously decarboxylates to form acetone

How is protein kinase A (cAMP-dependent protein kinase) activated by cAMP?

-Four cAMP subunits bind to PKA regulatory subunits inducing conformational changes -This releases two catalytic (active) subunits

Inner Mitochondrial Membrane (IMM)

-Freely permeable to only: O2, CO2, and H2O -Transport proteins exist for everything else - > 15 fold larger area than OMM due to invaginations called cristae - 75% protein by mass -Contains all the enzymes of electron transport and Ox-phosin

Mode 1: Much more ribose 5 Phosphate than NADPH is required

-Full reaction written in the image - 5G6P + ATP --> 6ribose 5-phosphate + ADP + 2H+

What is GTPase activity? What is the role of the GTPase function of heterotrimeric G-protein? why does the GTPase activity need to be slow?

-G-protein acts as a very slow GTPase hydrolyzing GTP to GDP. -The GTPase acts as an "egg timer" the enzyme/protein can work until GTP is finally hydrolyzed to GDP -If GTPase activity was too fast then the protein would not be able to do anything

How is Adenylate Cyclase activated by hormone

-GPCR: heterotrimeric G-protein coupled receptor -Roughly 800 in genome -1/3 of all drugs target the GPCR -receptor g-protein complex -Hormone receptor changes upon hormone binding -Hydrolyze GTP --> GDP -G-protein is a slow GTPase -activates adenylate cyclase to make cAMP -The GTPase activity of G-protein functions like an egg timer. -As soon as GTPis --> GDPi AC is inactive

how is the signal from glucagon amplified in the cell?

-Glucagon interacts with its receptor causing G protein conformational changes -This activates adenylate cyclase which produces a large number of cAMP from ATP which then amplifies the signal

what is glucagon, where is it made, and what is its effect?

-Glucagon is a hormonal peptide produced by pancreatic a cells. -It signals low blood sugar through hormonal regulation described above involving G protein coupled receptors. -Glucagon triggers a cAMP cascade that activates fructose biphosphatase 2 (FBPase 2) and inactivates PFK2. This activates gluconeogenesis and inactivates glycolysis.

Bacteria in gut

-Glucose --> H2 + CH4 + High fructose corn syrup -55% glucose 45% fructose -HFS (high fructose corn syrup) is a bacterial enzyme and bypasses pfk regulation -lactose fermentation mech. and HFS reaction tree shown in picture

Glycogen Structure

-Glucose linked by a(1 --> 4) linkages -a-D-glucose has an OH down at carbon 1 -opposite side as CH2OH -b-D-glucose has an OH up at carbon 1 -Same side as CH2OH -Glycogen grows and degrades from the nonreducing end (side with carbon 4 prominant) -Reducing end has carbon 1 and OH prominant -Crosslinks at a(1 --> 6) linkages: -Branches -Tollens reagent (ag) reacts with the a(1-->6) linkages

Amino Acid transport Proteins:

-Glutamate, aspartate: Glu/Asp carrier (antiporter)

Glucose-6-Phosphate Dehydrogenase plays a key role in the protection against Reactive Oxygen Species

-Glutathione (GSH) helps prevent damage by ROS generated over the course of metabolism -Oxidized glutathione (GSSG) is converted into reduced glutathione by NADPH -NADPH generated by G6P dehydrogenase -Glucose 6 Phosphate deficiency causes drug induced hemolytic anemia

What is glutathione and why is it necessary? How is it related to PPP?

-Glutathione is antioxidant found in plants -Useful in reducing the oxidative stress on a cell -Reduces H2O2 to H2O -The NADH produced by the PPP is needed to reduce GSSG (two glutathiones joined via a disulfide bridge) to glutathione (active form) -Useful in cells that undergoe a lot of oxidative stress (e.g. red blood cells)

Primer for Glycogen Synthesis

-Glycogen Synthase: how does it get started -requires a primer to initiate glycogen chain -Glycogenin (~37 kDa) -primer function -Intrinsic glucosyl transferase activity -Linkage of C1 to Tyr194

Diagram of Phosphorylase a and b Structure

-Glycogen binding site is highlighted in pink -Green highlighted loop leaves the active site open in phosphorylase a (R) and covers the active site in phosphorylase b (T)

Glucagon secretion is stimulated when blood glucose concentration decreases. Select all of the carbohydrate pathways that glucagon stimulates in the liver. -Glycogen breakdown (glycogenoysis) -Gluconeogenesis -Glycolysis -Glycogen synthesis (glycogenesis) -Glucose uptake

-Glycogen breakdown -Gluconeogenesis

Epinephrine is released in response to stress, and is a fight or flight hormone. Select all metabolic pathways that epinephrine stimulates. -Glycogen breakdown (glycogenolysis) in the liver and muscle -Gluconeogenesis in the liver -Glycolysis in muscle -Lipolysis in adipose tissue -Glycogen synthesis (glycogenesis)

-Glycogen breakdown (glycogenolysis) in the liver and muscle -Gluconeogenesis in the liver -Glycolysis in muscle -Lipolysis in adipose tissue

Select the results that occur from having few or no a-1,6 linkages in glycogen

-Glycogen degradation would slow down -Maintaining proper blood sugar levels would be more difficult

Fed State Favors Glycogen Synthesis and Storage

-Glycogen is a glucose polymer -Glycogen is most abundant in the liver and muscle -Its function varies according to the tissue its located in -Archea, bacteria, and eukaryotes all use glycogen -Plants use starch

Phosphorylase Mechanism

-Glycogen is the R group -A bound HPO42- donates a proton to the C4 oxygen of the departing glycosyl group -This is favored by the transfer of a proton from the protonated phosphate group of the bound PLP group -The carbonium ion and and glucose 1 phosphate combine to form glucose 1 phosphate -Mechanism is much clearer in the image -Glycogen phosphorylase removes >1 glucose 1 P while remaining bound -Schiff bases: primary amine + aldehyde

Glycogen in Skeletal Muscle

-Glycogen is utilized for ATP synthesis --> muscle contraction -Storage of glycogen, degradation to G6P consumption in glycolysis -Fed state: produces glycogen -Fasting state: produces ATP from glycogen (lactate byproducts)

Adrenalin / epinephrin, glucagon, insulin: where are they made, where do they act, and what effect do they have on glycogen metabolism?

-Glycogen metabolism shown in image -Organ: -where messengers released -stimuli -messenger -receptors -Vascular: -pituitary -Stress, low blood sugar -Vasopressin -a adrenergic receptors Liver: -Pancreas -Low blood glucose -glucagon -glucagon receptors Muscle/liver: -adrenal medulla -stress -epinephrine -epinephrine receptors Nervous system: -junction -nerve impulse -acetylcholine -Muscarinic receptors

Glycogen Phosphorylase Diagram

-Glycogen phosphorylase is bound to granules -Dimer -Glycogen binding site is 30 A ~5-6 glucose units big allowing it to phosphorolyze several glucose before rebinding -Each catalytic site includes a pyridoxal phosphate (PLP) group, linked to lysine 680 of the enzyme -The binding site for the phosphate (Pi) substrate is shown -The catalytic site lies between the C terminal domain and the glycogen binding site

what would the net outcome be if glycogen breakdown and synthesis were active at the same time?

-Glycogen synthesis and breakdown would occur at the same time. -Overall nothing would happen and body would just waste energy -"futile cycle"

Glycogen in the brain

-Glycogen utilized as a fuel source for G6P -Emergency supply of Glucose -Hypoglycemia (low blood glucose) -Hypoxia (low O2) -Majority of glucose is from the liver -Need a way to signal from the brain to the liver -Hormonal control (travels in the bloodstream)

Glycogen Synthase

-Glycogen(n) + UDP-Glucose --> glycogen(n+1) + glucose -Phosphorylating glycogen synthase: -Many glycogen synthase kinases -Protein phosphatase 1 (PP1) -Phosphorylated form inactive -Glycogen Synthase Kinases: -All kinases that activate phosphorylase deactivate synthase -cAMP dependent protein kinase: PKA inhibits synthase and activates phosphorylase kinase -phosphorylase kinase: inhibits synthase -Ca2+-calmodulin dependent protein kinase

familiarize yourself with glycogen storage diseases (Table 21-2) even though we didnt talk about it. think about how the clinical features might be explained.

-Google if need be -not worth it to list all explanations here

Photosystem I Half Reactions

-Half reactions in the image on the right -p700(red) --> p700(ox) is super unfavorable -p700(red) -hv-> p700*(red) super favorable

P680 Reduction

-Half reactions shown in the top left of the image -Mediated by the OEC a Mn protein complex bound to PSII -In the cycle 4e- go in the cycle and 4e- enter in the last step -The OEC must store 4e- from H2O and release one by one

Hexokinase inhibition

-Hexokinase isoforms are tissue specific -Hxk I, II, III: inhibited by G6P (product inhibition) -Hxk II: upregulated in cancer -Hxk IV: unique kinetic properties (liver/ pancreas) -Has a high Km for glucose since it has to send out glucose to other organs and G6P cannot leave the cell -Hxk are not good for flux regulation -Glucose + ATP --> G6P + ADP used for storage (glycogen)

Effects of Regulatory Compounds on Gluconeogenesis and Glycolysis

-High AMP: -Energy state low -Favors glycolysis -High citrate: -TCA low -Favors gluconeogenesis F 2,6 biphosphate: -Favors glycolysis -low energy elevated by AMP -Low blood glucose lowered by glucagon (through cAMP) -Glucagon: peptidehormone -signals low blood sugar

Allostery Defined by MWC model (T<-->R) Cont.

-High Energy State: -ATP UP, Glycogen Phosphorylase: inactive (T) -G6P UP, Glycogen Synthase: active (R) -AMP DOWN, Favors net glycogen synthesis -Low Energy State: -ATP DOWN, Glycogen Phosphorylase: active (R) -G6P DOWN, Glycogen Synthase: inactive (T) -AMP UP, favors glycogen breakdown -We need hormonal control to control organs

Response of PDH complex to the energy charge

-High Energy inhibitors: NADH, Acetyl CoA, ATP -Low Energy inhibitors: ADP, pyruvate -Inhibits PDH kinase

Glycogen Synthesis Overview:

-High blood glucose (1) glucose 6P <--phosphoglucomutase--> glucose 1-P -Same enzyme for degradation (2) glucose 1 P + UTP -UDP-glucose pyrophosphorylase-> UDP-glucose + PPi -PPi = 2Pi -activated form of glucose (3) UDP-glucose + glycogen(n) -glycogen synthase-> UDP + glycogen(n+1) -elevated energy status (4) glycogen branching enzyme -Steps 2-4 are unique to this synthesis -Overall strategy: activate glucose by forming a high energy sugar nucleotide

how is the amount of F 2,6 bisphosphate determined / regulated?

-High in fed state and low in starved state -F 2,6 biphosphate is allosteric effector of PFK whereas F 6 phosphate is not. -F 2,6 biphosphate stimulates PFK (glycolysis) and inhibits F 1,6 biphosphatase (gluconeogenesis) -When blood sugar level is low F 2,6 biphosphate loses a phosphoryl group to form F 6 phosphate. -PFK2 forms F 2,6 biphosphate -Fructose biphosphatase 2 (FBPase 2) forms F 6 phosphate -Both enzymes on the same polypeptide chain... Bifunctional enzyme

(2) Covalent Modification by Protein Phosphorylation

-Highly regulated: kinase kinase -Glycogen phosphorylase: -phosphorylase b (inactive) --> phosphorylase a (active) through phosphorylase kinase -Phosphorylase a (active) --> phosphorylase b (inactive) through Pi protein phosphatase 1 (PP1) -phosphorylase a frees up glucose -pretty non-specific (not that highly regulated) -reversible modification -Independent control -Hormonal regulation -Phos a: Phos b ratio is determined by relative activities of kinase vs phosphatase

E3: Dihydrolipoyl dehydrogenase (part 4/Ping Pong Mech.)

-Hydride transfer occurs between FADH- and NAD+ making FAD and NADH -E3 follows the ping pong mech. shown in the image

E2: Dihydrolipoyl transacetylase

-Hydroxyl-TPP reacts with lipoamide which changes conformation to dihydrolipoamide complex. -This forms a tetragonal intermediate which makes TPP and good leaving group -Results in a high energy thioester bond that reacts with coenzyme A -1st half of mechanism shown in the picture

Why metal cofactors are needed

-In biology e- and H+ move together (hydride) so we need metals (Fe2+/Fe3+) to only trade e-

why does the breath of diabetics in crisis smell of acetone?

-In diabetes there is insufficient glucose uptake and elevated b oxidation/gluconeogenesis -In diabetes there are a large number of ketone bodies in the blood. -These ketone bodies are degraded to acetoacetate which then spontaneously decarboxylates to form acetone -The acetone can then be smelled in the breath of someone with diabetes

Metabolic Interrelationships between the brain, muscle, adipose tissue, liver, and kidneys

-In diagram -G6P Fates: -Lactate from pyruvate through glycolysis and subsequent usage in brain and muscle -NADPH through pentose phosphate pathway -To glucose through the liver and released into the blood for other tissues -Pink arrows indicate pathways that predominate in the well fed state when glucose, fatty acids, and amino acids are directly available from the intestines

steps of FA synthase, and movement of the growing FA chain from ACP to a cysteine on the enzyme and back.

-In image

steps of FA synthase, and movement of the growing FA chain from ACP to a cysteine on the enzyme and back. Cont.

-In image

steps of FA synthase, and movement of the growing FA chain from ACP to a cysteine on the enzyme and back. Cont. Cont.

-In image

mechanism of thiolysis (alpha-beta cleavage)

-In image -Uses coASH

mechanism of thiolysis (alpha-beta cleavage) Cont.

-In image again

Table of Gluconeogenesis Reactions

-In image on the right

Components of the mitochondrial ETC

-In image on the right -D: e- donor -A: e- acceptor

STEP 2: PEP Carboxykinase (PEPCK)

-In mitochondria but mostly in cytosol -Driven by decarboxylation -Driven by GTP hydrolysis -Occurs often in the cytosol

Reciprocal regulation

-In the cell one pathway is relatively inactive whereas the other is highly active -Glycolysis will predominate when glucose is abundant -Gluconeogenesis will be highly active when glucose is scarce -Occurs in gluconeogensis and glycolysis -interconversion of 1,6-biphosphate and fructose 6-phosphate (2 enzymes) -interconversion of phosphophenolpyruvate and pyruvate -When both pathways are activated there is waste

(1) Activation of Fatty Acids

-In the cytoplasm -Fatty acid is activated by acetyl CoA synthase to form fatty acyl CoA -has a tetrahedral intermediate -Fatty acyl CoA has high energy bonds -Fatty acyl CoA is the activated form of fatty acids

Monod-Wyman-Changeux model of allostery Cont.

-In vitro ATP inhibits Phosphofructokinase (PFK1) - but in muscle ATP: 4 mM, ADP: 0.4 mM, and AMP 0.1 mM -Adenylate kinase: ADP + ADP --> ATP + AMP -PFK1 responds to a small depletion of ATP through a larger increase in AMP -PFK1 activity changes by 120x -[ATP] is buffered to 10x variation in [ATP]

Ketoacidosis is a potentially life‑threatening condition that can occur when there is inadequate cellular glucose uptake, such as in uncontrolled diabetes. Order the steps that would lead to the development of ketoacidosis.

-Inadequate -Glucose metabolism decreases -Ketone bodies are produced in the liver -Ketone bodies are used as fuel for tissues -Ketone bodies accumulate in the blood, causing the pH to decrease -Ketoacidosis

Which of the following stromal changes occur in response to light that regulate the calvin cycle?

-Increase in pH -Increase in the levels of Mg2+ -Increase the amounts of reduced ferredoxin

liver glucokinase (isoform)

-Induced fit closure of active site upon glucose binding -Brings glucose c6 in close proximity to ATP gamma P -Excludes from active site -Reaction tree shown in the left of the picture

Insulin Receptor Signaling

-Insulin stimulates glycogen by inactivating glycogen synthase kinase -ISPK: insulin stimulated protein kinases Insulin receptor: -Receptor consists of 2 units each with an a and b subunit -2 a units on the outside of the cell come together and form the insulin binding site -b units lie on the inside of the cell and include a protein kinase domain

Hormonal Control of Fructose 2,6 Biphosphatase

-Integration of signals from other organs -if no insulin receptor then you are oblivious to insulin -(i.e. liver cells) -Pancreas produces a cells under low blood glucose (fasting) which interact with glucagon receptors -pancreas produces b cells under high blood glucose (fed) which interact with insulin receptors -Hormonal controls is mediated through G-proteins -(cascade effects) -Secondary messenger amplification -cAMP dependent protein kinase A (cAK) is roughly PKA -Catalytic substrates (C) are bound to regulatory substrates (R) and become active when separated

Glycogen Synthase Cont.

-Interlaced regulatory pathways -allosteric (dense energy status of cell) -hormonal regulation to integrate signals throughout the body -opposite to phosphatase

What are the biosynthetic roles of the TCA cycle? What does cataplerotic and anaplerotic mean?

-Intermediates are drawn off for biosynthesis when the energy needs of the cell are met. -Intermediates are replenished by the formation of oxaloacetate from pyruvate and other metabolites. -anaplerotic: reactions are when CAC intermediates are generated from other metabolites. -cataplerotic: reactions are when CAC intermediates are depleted for anabolicsynthesis

Cytosol

-Intermembrane space (IMS) -Contains enzymes of glycolysis and some citric acid cycle enzymes. -pyruvate and NADH must enter the mitochondria from the cytosol -pyruvate into mitochondria and Acetyl CoA out of the mitochondria

how is cAMP made, why is this favorable, and how is it removed?

-Is produced from ATP by adenylate cyclase (on G protein) -Glucagon trigger a cAMP cascade -cAMP is degraded by phosphodiesterase (PDE) to form AMP (inactive) -Glucagon binds favorably to the its receptor on the G protein. This causes conformational changes that activate adenylate cyclase to form cAMP

If Complex II (succinate DH) doesn't transport protons, what happens to the energy from succinate oxidation?

-It is transferred to complex 3 through CoQ

How does Coenzyme Q move between complexes?

-It moves freely through in the membrane because of its isoprene tail

Other sources of carbon for gluconeogenesis

-Lactate (from muscle) -Amino Acids -glycerol ("BB" from fat)

Phototroph

-Light energy to feed self - ~60% are bacteria (cyano) - ~2% of light is absorbed

Light and Dark Reaction comparisons

-Light reactions make NADPH and ATP -Dark reactions use NADPH to fix CO2 --> glucose -RuBisCo: CO2 fixation -glucogenesis -Pentose Phosphate Pathway

Lipase Figure

-Lipases convert triacylglycerides --> diacylglycerides --> monoacylglycerides

Lipoamide/dihydrolipoamide

-Lipoamide: oxidized -dihydrolipoamide: reduced -specifically the S-S bonds -These are both the active portions of E2 -dihydrolipoamide has a more positive standard reduction potential (lower case epsilon) than NADH -more positive standard reduction potential the higher affinity for e-

Pentose Phosphate Pathway (PPP) Summary

-List of pathways that require NADPH is in the top left of the image -Glyceraldehyde 3 phosphate is a Sciff base in the transaldolase step -Thiamine pyrophosphate TPP forms a carbanion in the transketolase step -Transaldolase: Steps listed in the left of the image -Transketolase: Steps listed in the right of the image

Fuel Reserves in a 70 Kg Human

-Liver exports most of is glucose and glycogen stores -Muscle has a large amount of all stores (mostly protein) -Adipose tissue is almost exclusively triglycerides -Brain has only minimal fuel reserves (a tiny amount of glycogen and glucose)

(3) Glucose 6 Phosphatase

-Liver not muscle -Favorable dG0' ~= -14 kJ/mol

Observation of ATP driven rotation in ATP synthase

-Long actin arm with fluorescent label on the end attached to gamma subunit. -Counter clockwise rotation of the gamma subunit -a3b3 is tacked down peripheral arm -gamma subunit in the a3b3 center (charge neutral) -As gamma rotates it induces conformational changes in directionality L --> T --> O (loose --> tense --> open) -This drives directionality of ATP synthesis Differences in ATP concentration: -At 200 nM of ATP: -Reaction goes backward -ATP + H2O --> ADP + Pi - T --> L --> O -At 20 nM of ATP: -Occasionally goes in the reverse direction - O --> L --> T

Hormonal Control:

-Low blood glucose makes more glucagon which amplifies the signal -cAMP is then produced and amplifies the signal further to fructose 6 biphosphatase -PFK is less activated so less glycolysis -Less inhibition of FBP 2 leads to more gluconeogenesis -Gluconeogenesis mainly takes place in the liver -specialized compartments: -Glucose 6 p'tase -Glucagon receptors -Organs without receptors will not respond

Regulation of glycogen synthase

-Many glycogen synthase kinases phosphorylate (inactivate) glycogen synthase -Protein phosphatase-1 dephosphorylate (activate) glycogen synthase

The Electron-Transfer Potential of an Electron

-Measured as redox potential (Eo') -Measure of a molecule's tendency to donate or accept electrons -Delta Go' = -nFdeltaEo' (n is the number of electrons transferred and F is the faraday constant)

Heterotrimeric G-protein and how it activates adenylate cyclase in response to hormone binding to GPCR.

-Mechanism as depicted above with adenylate cyclase acting as the enzyme making cAMP through hydrolysis

Phosphoglucose isomerase (PGI) Cont.

-Mechanism in enzyme -B: AA -B': LYS, HIS

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) Cont.

-Mechanism in enzyme -active site cysteine forms a high energy thioester in step 2 as well as a hydride transfer -Energy stored in high energy thioester -A high energy bond is formed gaining reducing power. -Step 4 (final step): orthophosphate attacks the thioester forming 1,3-BPG (product)

Complex IV: Cytochrome c oxidase Cont. x5

-Mechanism of Cyt. C in the image on the right -Reduction of O2 by Cyt. C oxidase makes a resonance stabilized radical -2 e- from Fe(II) --> Fe(III) -1 e- from tyrosine -1 e- from Cu(I) -replenished with 4e- from Cyt. C. -1-4 from 4 Cyt. C to restore the system

E1: Pyruvate dehydrogenase

-Mechanism shown in the picture -Hydroxyethyl-TPP: E1 product

Why are there no mitochondria in red blood cells?

-Mitochondria would use up all the oxygen that red blood cells have to transport

(1) Allostery described by the MWC model (T <--> R)

-Monod-Wyman-Changeux model -Protein can exist in at least two functional states: tense and relaxed. State is determined by thermal equilibrium -Binding of effectors only shifts the probability that the protein will be in one state or the other. -Effectors will not make the conformational change but shift towards the other conformation. Glycogen Phosphorylase: -glycogen(n+1) --> glycogen(n) + G1P -T(inactive) <--> R(active) -tense effectors: glucose, G1P, ATP -relaxed effectors: AMP Glycogen Synthase: -glycogen(n) + UDP-Glucose --> glycogen(n+1) + UDP -T(inactive) <--> R(active) -tense effectors(?): adenosine, caffeine -relaxed effectors: G6P -All effectors above are allosteric effectors

how does insulin act in response to high blood glucose level?

-More insulin is released

Ribulose Bisphosphate Carboxylase Oxygenase (RuBisCo)

-Most abundant protein on the plant -5Kg/person -Catalyzes CO2 fixation (~560 kDa) -Mechanism shown in the right of the diagram -Makes enough e- to make glucose (final product is GAP)

Gluconeogenesis Pathway

-Mostly in liver, kidney -Liver only stores ~12th worth of glucose -Gluconeogenesis goes up in the pathway on the rileft of the image and glycolysis goes down in the pathway

where is glycogen stored (organ / where in the cell)?

-Mostly stored in the liver and muscles -Glycogen is stored as granules in the cytoplasm -Stored as a cross linked polymer

(III) Isocitrate Dehydrogenase

-NADH (product inhibition) -ATP (allosteric inhibitor) -ADP (allosteric activator) -Ca2+ (allosteric activator)

(IV) alphaKetoglutarate Dehydrogenase

-NADH (product inhibition) -Succinyl CoA (product inhibition) -Ca2+ (allosteric activator)

Malate/a-ketoglutarate carrier and glycerophosphate shuttle Cont. Cont.

-NADH + 1/2O2 + H+ <--> NAD+ + H2O -dEo' = 1.13 V -dGo' = 218 kJ/mol (super favorable)

Complex I: NADH: ubiquinone (CoQ) oxidoreductase Cont.

-NADH + Q + 5H+(matrix) --> NAD+ + QH2 + 4H+(intermembrane space) -vectorial proton pump -Ubiquinone: -Coenzyme Q (CoQ/Q/Q10) -oxidized quinone -"greasy" -Ubisemiquinone: -Coenzyme QH. -radical -semiquinone -Ubiquinol: -Coenzyme QH2 -reduced -active form -hydroquinone -NAD+ only deals in H+ 2e- -CoQ can do 1 or 2 e- + H+ -Fe2+/Fe3+ only deals in 1 e-

Complex I: NADH: ubiquinone (CoQ) oxidoreductase

-NADH <--> NAD+ + 2e- + H+ (0.315 V) -CoQ (ubiquinone) + 2e- +2H+ <--> CoQH2 (ubiquinol) (0.045 V) - dEo' = 0.036 V - dGo' = -69.5 kJ/mol (Formally can make ATP)

What can block components of the ETC?

-NADH-Q Oxidoreductase (complex I): -rotenone (fish poison) -amytal (barbiturate) -Q-cytochrome C Oxidoreductase (Complex III): -antimycin A (fungicide) -Cytochrome C Oxidase (Complex IV): -CN- -N3- -CO

Plant Photosystem (Complex IV)

-NADP+ Reductase: - e- from p700 chla reduce NADP+ to NADPH -Mediated by ferrodoxin -FAD is a cofactor

To answer this question, you may reference the Metabolic Map. Which molecules directly participate in fatty acid synthesis by acting as energy sources? Ethanol consumption inhibits gluconeogenesis by causing an increase in the concentration of NADH. NADH is a substrate for malate dehydrogenase, the enzyme that produces malate. An accumulation of malate favors the activity of malic enzyme. Thus, an increase in NADH concentration increases the enzymatic activity of malic enzyme.

-NADPH and ATP -Increase in NADPH concentration, stimulates fatty acid synthesis, ethanol consumption in a fasting state increases the risk of hypoglycemia

1) NADPH is not interchangeable for NADH

-NADPH and NADH are not metabolically interchangeable (usually) -Remember NADPH is just an ADP with a nicotinamide attached -NADH: -catabolic pathways -carbon oxidation to CO2 -ATP synthesis -NADPH: -Synthesis of: FAs, AAs, nucleotides, glutathione (reducing agent... detoxify reactive oxygen species), degradation of heme. -[NAD+]/[NADH] = 10^3 (favors oxidation) -[NADP+]/[NADPH] = 0.01 (favors metabolic reduction)

is food that contains mono- and diacylglycerols really 'low fat', 'low calory'?

-No a food containing mono and diacylglycerols still has all the energy associated with those fats -The "low fat" moniker is just due to the definition of fats not including mono and diacylglycerols

What is the oxygen evolving complex? What is the importance of Mn ions?

-OEC -Water splitting complex -Water oxidizing enzyme involved in the photo-oxidation of water during the light reactions of photosynthesis -Mn holds the electrons as they are stripped from water to make O2 -Electrons are removed four at a time but can only be placed onto Mn one by one

Draw the mechanism for the Fenton reaction. How is oxygen converted into a hydroxyl radical? Include arrows.

-OH. forms covalent adducts with DNA, protein, and lipids. -OH. (hydroxyl radical): "molecular scissors" -Through the fenton reaction forming first a super oxide then H2O2. H2O2 is then split into two OH. radicals by Fe2+.

Complex II: Succinate dehydrogenase Cont. x2

-OH. forms covalent adducts with proteins, lipids, and DNA -In complex 2 FADH is the entry point of e- -Does not pump protons but instead gives them to Q to form QH2 -fenton reaction: "biological scissors"

Steps that drive the TCA Cycle

-Occur early in the TCA cycle

(3) Alpha Beta Bond Cleavage

-Occurs in the mitochondria -ETF: electron transferring flavoprotein -membrane bound -Specific enzymes for fatty acid lengths: -long: 12-18 -medium: 6-12 -short: 4-6 -alkane --> keto: -Similar to TCA cycle (succinate --> oxaloacetate) -Different enzymes similar chemistry

Step 1: Pyruvate Carboxylase

-Occurs in the mitochondria -Two phases Phase 1: -Hook P onto C --> carboxy phosphate --> carboxy biotin -Biotin = vitamin H Phase 2: -carboxybiotin --> OAA

Allosteric control of fructose 1,6 biphosphate

-Occurs through the control of fructose 1,6-biphosphate (FBP) by AMP, citrate, and fructose 2,6 biphosphate -Gluconeogenesis and glycolysis are linked through these substrates -PFK-2: phosphofructokinase 2 -FBP-2: Fructose biphosphatase 2 -Gluconeogenesis: F 1,6 BP to F6P -Glycolysis: F6P to F 1,6 BP

Reciprocal regulation of F1,6 bisphosphatase and Phoshopfructokinase by AMP, Citrate, and F2,6 bisphosphate

-Occurs through the control of fructose 1,6-biphosphate (FBP) by AMP, citrate, and fructose 2,6 biphosphate -Gluconeogenesis and glycolysis are linked through these substrates -PFK-2: phosphofructokinase 2 -FBP-2: Fructose biphosphatase 2 -Gluconeogenesis: F 1,6 BP to F6P -Glycolysis: F6P to F 1,6 BP Low AMP: Glycolysis (Phosphofructokinase) High AMP: Gluconeogenesis (F 1,6 biphosphatase) Low Citrate: Glycolysis (Phosphofructokinase) High Citrate: Gluconeogenesis (F 1,6 biphosphatase) Low F 1,6 bP: gluconeogenesis (F 1,6 biphosphatase) High F 1,6 bP: glycolysis (Phosphofructokinase)

Issues with unsaturated FA degradation. what if the double bond is at an even-numbered carbon? what has to happen if it is on an odd-numbered carbon?

-Odd chain fatty acids: -yields 1 propionyl CoA -B oxidation alone cannot degrade unsaturated fatty acids -Unsaturated fatty acids with an odd number of double bonds need only isomerase -Unsaturated fatty acids with an even number of double bonds need both isomerase and reductase

Why is sucrose-octaester (olestra) a low-calory fat substitute?

-Olestra cannot be digested yet tastes and feels like fat. -Therefore olestra has very little to no calories associated with it

Red Blood cells:

-Only eat glucose -Do NOT have mitochondria

UDP-Glucose Pyrophosphorylase Cont.

-Overall -19 kJ/mol -First reaction is non-favorable (dG0' = 0) -Second reaction is favorable (dG0' = -19) -When the two reactions are coupled the overall reaction is favorable

Dark Reactions (Calvin Cycle)

-Overall Calvin Cycle in the image on the right -Rubisco enters stage 1 (fixation) -In stage 2 GAPDH: Pi + NAD+ --> NADH + ATP -Learning Goals: -explain the function of the calvin cycle -Describe how light reactions and the calvin cycle are coordinated

Energetics of Gluconeogenesis

-Overall Gluconeogenesis shown in image -Enzymes of gluconeogenesis also shown -Mitochondria: ATP>>ADP -dGo' values written below -Overall 6 P are spent 1) Pc/PEPCK vs pyruvate kinase: (1 vs -23) 2) FBPt'ase vs PFK: (-16 vs -17) 3) G6Pt'ase vs hexokinase (-14 vs -21)

Complex IV: Cytochrome c oxidase (Cox) Cont.

-Oxidase: O2 is reduced -4 Cyt. C(red) + 8H+(matrix) + O2 --> 4Cyt. C(ox) + 2H2O + 4H+(intermembrane space) -vectorial transport of 4H+ -Prosthetic groups: -Two Hemes: -Heme a -Heme a3 -Two copper centers: -CuA -CuB -They form the peroxide bridge (bind to O2) -In square planar or tetrahedral form -CuA: 2 copper center: Cu(1)-Cu(1) -Loses an electron to form Cu(II)-Cu(I) or Cu(1.5)-Cu(1.5) (delocalized) -Similar to [Fe-S]

Oxidative Stage of PPP

-Oxidation of Carbon to CO2 -Reduction of NADP+ --> NADPH +H+ a) Glucose 6P dehydrogenase: -C1 aldehyde of G6P is oxidized to acid -2e- --> 2NADPH b) 6 Phosphoglucose lactonase: -H2O ring opening -C3 alcohol further oxidized to C2 ketone -2e- --> NADPH -Overall: G-6-P --> --> CO2 + Ru5p (ribulose 5 phosphate) + 4e- 4e- + 2NADP+ --> 2NADPH + 2H+

how are odd chain fatty acids synthesized (and remember how they are broken down, as well!) Cont.

-Oxidized (metabolized) with 3 enzymes: (1) propionyl CoA carboxylase (2) Methylmalonyl-CoA epimerase (3) Methylmalonyl-CoA mutase

Pyruvate Dehydrogenase (PDH)

-PDH is regulated by post translational modification (PTM) -Regulated by substrates of the TCA cycle: OA, Acetyl CoA -Regulated by the products of the TCA cycle: NADH

Table of Regulators of Gluconeogenic Enzyme Activity

-PFK 2 and FBPase 2 are bifunctional enzymes

Protein Phosphatase 1 (PP1)

-PP1 active: -glycogen synthase on -glycogen phosphorylase off -Glucose storage favored -Made specifically by interchange? proteins (~2200) -Quite non-specific... many targets -Removes P signals -Bound to glycogen particles via G subunit (~160 kDa) -G is phosphorylated at 2 serines (site 1 and site 2) by cAMP dependent protein kinase -G is phosphorylated at site 1 by "insulin-stimulated protein kinase" (ISPK) -ISPK as stimulates phosphodiesterase which destroys cAMP -ISPK also activates glycogen synthesis and inhibits glycogen degradation -More glucose uptake -Insulin stimulates glycogen synthesis by activating a signal transduction pathway that results in the phosphorylation and inactivation of glycogen synthase kinase. -PP1 subsequently dephosphorylates glycogen synthase (generates the active a form) -Insulin also facilitates glycogen synthesis by increasing the number of glucose transporters (GLUT4) in the plasma membrane... increases the uptake of glucose

regulation of protein phosphatase through G-protein.

-PP1 is bound to glycogen via G subunits -G subunit is phosphorylated at two serines (site 1 and 2) by cAMP dependent protein kinase -G subunit is phosphorylated at site 1 by ISPK

Plant Photosystem (5 complexes)

-PSI: light -PSII: light -Cyt. bf6: complex III (ETC) -plastoquinone: mobile e- carrier -plastocyanin: mobile e- carrier -3 transmembrane protein complexes (1, 2, and 3) linked by mobile e- carriers (plastoquinol and plastocyanin) -proton translocating ATP synthase (like us) -Light driven transport of e- from H2O to NADP form NADPH -Fd: ferrodoxin -OEC: oxygen evolving complex

Segregation of Photosystem I and Photosystem II

-PSII in stacked region to keep away from PSI -If PSII and PSI were close II680 would be attenuated for I700 -In the image the distribution of stacked (grana) to the unstacked (stroma exposed) regions of the Tk membrane is shown

Photooxidation Charge Figure

-PSII: -p680 chla dimer -pheophytin a (chla, -Mg) -plastoquinones QA, QB -2Q + 2H2O -hv-> O2 + 2QH2 -p680*: excited p680 -formally transfers e- from water to photosystem 1 in ETC

Reduced Glutathione (GSH)

-Peroxides react with GSH to form alcohol and GSSG essentially terminating the ROS

Signal Integration Diagram

-Phosphodiesterase also acts in the same way as insulin stimulated protein kinase Organ: -where messengers released -stimuli -messenger -receptors -Vascular: -pituitary -Stress, low blood sugar -Vasopressin -a adrenergic receptors Liver: -Pancreas -Low blood glucose -glucagon -glucagon receptors Muscle/liver: -adrenal medulla -stress -epinephrine -epinephrine receptors Nervous system: -junction -nerve impulse -acetylcholine -Muscarinic receptors

Identify the enzymes that are required for the synthesis of a glycogen particle starting from glucose 6-phosphate

-Phosphoglucomutase -Pyrophosphatase -Glycogenin

Phosphoglycerate mutase Cont.

-Phosphohistidine at catalytic site -2,3 BPG in blood helps release O2 (think Biochem) -Binds to deoxy Hb and weakly to Oxy Hb -genetic disorders in glycolysis alter capacity for blood to transport O2. -Fetal Hb lower affinity for 2,3 BPG competes for O2 from maternal Hb

(4) Debranching Enzyme

-Phosphorylase catalyzes processive cleavage to 4-5 residue from the branch point a(1-->6) -Has 2 active sites an a-1,4 glucanotransferase (GT) and an a-1,6 glucosidase (GC) -bifunctional enzyme -When the limit branch is reached a-1,4 glucanotransferase (GT) transfers the "stub" to a nonreducing end - a-1,6 glucosidase (GC) thens cleaves the "stub" forming glucose and a debranched glycogen

Simplified Plant Photosystem

-Photon absorption by PSII and PSI powers e- flow from water to NADP+ to make NADPH -This is used to make complex CHO, [H+] gradient (to make ATP), and O2 (waste product). -Oxygen evolving complex (OEC) and water evolving complex (WOC) are the same thing -e- is knocked loose and replaced by the e- from water

Diagram comparing Photosynthesis with Oxidative Phosphorylation

-Photosynthesis: -high e- hv -replenished from H2O -Dumped onto NADP+ -Oxidative Phosphorylation: -e- from CHO --> O2 --> H2O -H+ gradient

Thioredoxin and the regulation of the Calvin Cycle

-Photosystem I generates reduced ferrodoxin and NADPH --> regulate enzymes of the Calvin Cycle -Ferroredoxin-thioredoxin reductase: activates key enzymes by reducing disulfide bonds that control their activity -Table of enzyme regulated by thioredoxin in the image on the right

Plant Photosystem (Complex III)

-Photosystem I: -Energy form LHC drives photooxidation of chla (p700) -chla+ reduced by e- transfer from Cyt. b6-F -Mediated by plastocyanin (Cu protein) -Pc: plastocyanin -Look to the image on the right for the reaction

Light Harvesting Center II (LHC II) from peas chloroplasts Cont.

-Pigments are suspended in different orientations to harvest light from many angles and many wavelengths -Protein is used to hold the pigments in place and facilitate energy transfer. -Side view of LHC II in image on the right

What is photorespiration, what causes it and why is it wasteful?

-Plants consume O2 and release CO2 -Caused by rubisco oxygenase activity -Organic carbon becomes CO2 without producing any energy rich compounds

Understand the importance of coupling reactions in the UDP-glucose pyrophosphorylase mechanism and what drives this reaction

-Powered by PPi to 2Pi (hydrolysis) -Group transfer and phosphoanhydride exchange

Why glucose?

-Primordial soup: could have been made from formaldehyde -High propensity to form: ring aldehydes, glycolysate proteins, can react amine groups

Overall glycogen Synthesis and Breakdown Cont.

-Problem futile cycle -solution: reciprocal regulation at glycogen phosphorylase and synthase Glucose utilization: -glycogen synthase: down -glycogen phosphorylase: up -fasting (liver) -exercising (muscle) Excess Glucose: -glycogen synthase: up -glycogen phosphorylase: down -Feeding Mechanisms: -allosteric effectors -Hormonal control -covalent modification through phosphorylation

ATP in information transfer

-Protein folding -organelle trafficking -signal transduction

Model of Photosystem

-Proton gradient from NADP+ reductase to power complex 5 (ATP synthase) -E = hv [680 um] [700 um]

Components of the Electron Transport Chain (ETC)

-Proton transport across the membrane In Image: -Top: The electrons donated by NADH and FADH2 are passed to electron carriers in the protein complexes -Right: Coenzyme Q -Left: Cytochrome C: carriers electrons from complex III to complex IV -Electrons flow down an energy gradient from NADH to O2 -Catalyzed by 4 protein complexes and the energy released is used to generate a proton gradient

Vectorial Transport in Cytochrome C Oxidase

-Protons bind through side chain protonations -protons move by relay (proton wire)

Gluconeogenesis Overview

-Pyruvate (lactate and glycerol), TCA intermediates, and amino acids are made into oxaloacetate (OAA) --> PEP --> glucose 1) Reversal of glycolysis: -Except: 4 steps hexokinase, phosphofructokinase, pyruvate kinase -glycolytic enzymes -far from equilibrium -Therefore independent regulation occurs 2) Conditions that favor gluconeogenesis inhibit glycolysis and vice versa -Therefore reciprocal regulation

Tricarboxylate Transporter (TC transporter)

-Pyruvate into the mitochondria Acetyl CoA out of the Mitochondria -How pyruvate gets into the mitochondria for use in the TCA cycle -How Acetyl CoA gets out of the mitochondria for fatty acid synthesis -NADPH: Needed for fatty acid synthesis (reducing power) -ATP citrate lyase makes Acetyl CoA from citrate to be used in fatty acid synthesis -Decarboxylation of BKeto acid

Glycolysis transport proteins:

-Pyruvate: pyruvate/H+ symporter -PEP (phosphorylated pyruvate): PEP transporter

Q cycle (Part of complex III function)

-Q Cycle: mech. for coupling electron transfer from QH2 to Cyt. C -The shuttling of electrons between ubiquinol and ubiquinone in the inner mitochondrial membrane as a part of Complex III's function -QH2 carries two electrons -Cyt. C carries one electron -In one cycle 4 protons are pumped out of the mitochondrial matrix and 2 additional ones are removed from the matrix -2QH2 + Q + 2Cyt. C(ox) + 2H+(matrix) --> 2Q + QH2 + Cyt. C(red) + 4H+(intermembrane space) -Q in reduced form pick up 2H+ -In first half of the cycle two electrons of a bound QH2 are transferred. -One to Cyt. C -One to bound Q in a second binding site to form the semiquinone radical ion Qe- -In the second half of the cycle a second QH2 gives up its electrons to complex III. -One to a 2nd Cyt. C -One to reduce Qe- to QH2

Q cycle (Part of complex III function) Cont. x2

-Q cycle is an electron transport cycle in complex III

Photosystem II and the Oxygen Evolving Complex (OEC)

-Q pool in top right of the graph -p680* is a strong oxidant that removes electrons from water -Occurs at the water-oxidizing complex (aka manganese center) of photosystem II -Four photons are required to generate one O2 -The four protons used to reduce the Q molecules come from the stroma -The four protons liberated from the water are released into the lumen

Channeling in multi-enzyme Complexes

-Rate enhancement: minimize distance between active sites -Specificity: minimize side reactions -E1, E2, and E3 sites in reaction shown

Regulation of Pentose Phosphate Pathway (PPP)

-Rate of the oxidative phase is determined by the level of NADP+ (most important regulatory factor) -Dehydrogenation of glucose 6 phosphate by glucose 6 phosphate dehydrogenase is the rate limiting step of the pathway

Prosthetic group of ACP

-Reaction driven by decarboxylation of malonyl ACP -Tail to head

Hexokinase Cont.

-Reaction is highly favorable -Gamma phosphate (ATP): Higher P-transfer potential than G6P -"energized glucose" -P traps glucose in the cell -Delta G" values of half reactions and overall reaction in the picture.

Standard Reduction Potentials of Some Biochemically Important Half-Reactions

-Reaction potentials in diagram -Strong oxidizing agent (large positive Eo') is a strong e- acceptor -Strong reducing agent (large negative Eo') is a strong e- donor

Respirasome Supercomplex

-Reactions catalyzed by respirasome super complex -Seen through cyro EM -Proton gradient from low H+ --> High H+ -

Anaplerotic Reactions

-Reactions that help replenish intermediates in the TCA cycle when their reserves are depleted. -TCA cycle intermediates regenerated from other metabolites

how is this outcome avoided?

-Reciprocal regulation at glycogen phosphorylase and synthase -In glycogen phosphorylase: -phosphorylation and dephosphorylation to form tense and relaxed forms -In glycogen synthase: -regulated through G6P

Regulation of Citric acid Cycle

-Regulated through 4 pathways -Regulated by two main ways: -Binding of metabolites -Covalent modification of enzymes

Multicyclic Cascades

-Results in amplification of signal -chemical -sensitivity -Divergence in pathways -Epinephrine and glucagon are hormones -1 enzyme can turn out many substrates -enzyme goes through cycles of glycogen --> G1P until glycogen concentration gets too low

Citrate Synthase: Cont.

-Reverse aldol condensation -Favorable due to Delta Go' = -31 Kj/mol thioester bond hydrolysis -Rate limiting step: Acetyl CoA binds after OA

Complex III: Coenzyme Q-cytochrome C Oxidoreductase (Cytochrome bc1 complex) Cont. x2

-Rieske [Fe-S] center coordinate Fe -has 2 His and 2 Cys -Cytochrome C (Cyt. C) picks up heme bl -Coenzyme Q-cytochrome C Oxidoreductase is a homodimer with each monomer consisting of 11 distinct polypeptide chains -Q stands for quinone -Electron transfer between quinones in the membrane and Cyt. C in the IMS is mediated by the 3 major prosthetic groups

C​alculate the standard free energy for translocating one mol of protons from the mitochondrial matrix to the intermembrane space, if the difference in pH across the inner mitochondrial is 0.75 (external side is more acidic), the difference in membrane potential is 100 mV (inside negative), and the temperature is 310 K.

-Same equation as the above question

how is cAMP-dependent protein kinase activated by cAMP (release of catalytic subunit through cAMP binding to regulatory subunit.

-Same process as with G proteins -4 cAMP residues bind to the 2 regulatory domains causing conformational change. This releases the 2 active catalytic subunits

Common Biological Fatty Acids Table

-Saturated fatty acids have no double bonds -Unsaturated have various number and position of double bonds -Cis: H are on the same side of the double bond -Trans: H are on opposite sides of the double bond

Overall Glycogen Synthesis and Breakdown

-Separate pathways, independent control, both directions are energetically favorable Overall: -glycogen(n+1) + H2O --> glycogen(n) + glucose (-18 kJ/mol) -Glucose + ATP + UTP + glycogen(n) + H2O --> glycogen(n+1) + ADP + UDP + 2Pi (-42 kJ/mol) -Overall: ATP + UTP + 2H2O --> ADP + UDP + 2Pi

PEP Carboxykinase (PEPCK) Cont.

-Separation of pyruvate carboxylase and PEPCK: -There is no direct transport of OAA -The formation of oxaloacetate by pyruvate carboxylase occurs in the mitochondria -OAA is reduced to malate and transported to the cytoplasm where it is reoxidized to OAA with the generation of cytoplasmic NADH -PEP is then synthesized from OAA by phosphoenolpyruvate kinase. -Decarboxylation often drives unfavorable reactions (i.e. TCA cycle, PPP, fatty acid synthesis)

Reaction catalyzed by 'malic enzyme', and fate of pyruvate and NADPH that is produced by it.

-Sequential activation of cytoplasmic malate dehydrogenase and malic enzyme -Malic enzyme catalyzes malate --> pyruvate + NADPH -pyruvate enter the mitochondria and then is converted to oxaloacetate by pyruvate carboxylase

Progressive alteration of the forms of the 3 active sites of ATP synthase

-Shown in bottom left of the image -No two subunits are every in the same conformation

Energy balance of making a fatty acid

-Shown in image

location of fatty acid synthesis and degradation (organ / in the cell)

-Shown in image

overview of energy utilization. what happens in the main organs during fed, fasting and starving fate? which metabolites are taken up / exported?

-Shown in image

Role of PLP in glycogen phosphorylase reaction, overall mechanism.

-Shown in image -schiff base -acts as a proton acceptor

Four Modes of PPP Reactions

-Shown in the image

Four modes of PPP Mechanisms

-Shown in the image -Mode 1: Needs ribose 5P (all carbons saved) -Mode 2: Ribose 5P and NADPH needed -Mode 3: Needs NADPH -remakes G6P (gluconeogenesis) -Mode 4: NADPH and ATP needed

Transketolase Mechanism

-Shown in the image -The activated glycoaldehyde in step 4 attacks the aldose substrate to form a new C-C bond

Transaldolase Mechanism

-Shown in the image along with listed steps

what are chylomicrons, and what is their architecture?

-Similar to HDL and LDL -Lipid protein aggregates -apoprotein, phospholipids, cholesterol, and cholesterol esters on the outside of the chylomicrons -Triglycerides on the inside -Essentially triglyceride transports in the blood and can be targeted to certain tissues -apoproteins can be recognized by cell receptors

specific acid-base catalysis

-Solvent acts on acid/base -HB+: lewis acid (e- acceptor) B: lewis base (e- donor)

Three Stages of the Calvin Cycle

-Stage 1 (fixation): Fixation of carbon by the carboxylation of ribose 1,5-bisphosphate -Stage 2 (reduction): The reduction of the fixed carbon to begin the synthesis of hexose (NADPH) -Stage 3 (regeneration): The regeneration of the starting compound ribulose 1,5-bisphosphate (similar to pentose phosphate)

(2) UDP-Glucose Pyrophosphorylase

-Step 1 is covered in glycogen degradation (above) -UTP: the three phosphates are labeled a, b, g from outside to inside -UDP-glucose: uridine diphosphate glucose -Group transfer of G1P for PPi on UMP -Phosphoanhydride exchange -Powered by PPi --> 2Pi -dG0' = -30 kJ/mol

During ketogenesis, the liver synthesizes ketone bodies that can be used as an energy source. Put the statements regarding ketogenesis in the correct order, beginning with a stimulus for ketogenesis.

-Stimulus -low blood glucose levels stimulate the breakdown of fatty acids to acetyl CoA -Two acetyl CoA condense to form the four carbon acetoacetyl-CoA -A condensation reaction with acetyl-CoA produces the 6 carbon HMG-CoA -HMG-CoA loses acetyl-CoA, forming a 4 carbon ketone body -3-Hydroxybutyrate and acetoacetate can cross the blood brain barrier to provide fuel for the brain.

Complex II: Succinate dehydrogenase Cont.

-Succinate + FAD --> fumarate + FADH2 -FADH2 + Q --> FAD + QH2 -Prosthetic groups: -FAD -3 [Fe-S] clusters -Heme b560 (Fe) -Cofactor arrangement: FAD --> [2Fe-2s] --> [4Fe-4S] --> [3Fe-4S] --> Q -Role of heme: -Not on direct path of e- transfer -Protects against reactive oxygen species (ROS) which are toxic -O2 --> HO2.- --> H2O2 -H2O2 + Fe2+ (fenton reaction) --> OH- + OH.+ Fe3+

Succinate to NAD+/FAD

-Succinate --> NAD+: -0.33 V -Succinate --> FAD: -0.031 V Not favorable

What are the purposes of the pentose phosphate pathway?

-Synthesis of NADPH and ribos 5 phosphate (R5P) -Synthesis of glycolytic intermediates

Standard reduction potentials for some biochemically important half-reactions

-Table in the image on the right

Model of E. Coli F1F0 ATPase

-The H+ wells channel through the aSubunit -With each turn one H+ flows into a well and one H+ flows out of the well ahead of it. -Essentially offset channels which (one in and one out) which the wells rotate between releasing an H+ every turn. -In subunit C (C ring) aspartic acid is the H+ acceptor

Diagram of the Core of the Water-Oxidizing Complex

-The absorption of photons by the reaction center generates a tyrosine radical which then extracts electrons from manganese ions -Mn can exist in 2+, 3+, 4+, 5+ -KoK cycle is depicted in the image on the right

bifunctional enzyme - which part of it is regulated by glucagon and thus by cAMP?

-The activity of either enzyme is reciprocally controlled by the phosphorylation of a serine residue. -In a fasting state glucose is scarce and glucagon rises -Glucagon triggers a cAMP cascade -Protein kinase A (PKA) phosphorylates that serine residue activating FBPase 2 and inhibiting PFK2

In photosystem II, P680 transfers an electron to pheophytin upon excitation by a photon. How is this electron replaced? Explain the challenges with this process

-The electron is replaced by the electrons resulting from the splitting of H2O into O2 from the OEC -P680 is only able to hold onto one electron whereas the splitting of water releases 4 electrons simultaneously so those electrons need to be held onto until they can be picked up by P680

Identify the true statements regarding a-1,6 linkages in glycogen

-The enzyme that forms a-1,6 linkages is catalyzed by a branching enzyme

Cyclic photophosphorylation diagram

-The flow of electrons through cytochrome bf pumps protons into the thylakoid lumen.

Gluconeogenesis Facts

-The generation of free glucose (final step of gluconeogenesis) only occurs in the liver -G6P is transported to the lumen of the endoplasmic reticulum -Glucose 6-phosphatase: an integral membrane on the inner surface of the ER catalyzes the the formation of glucose from G6P -In tissues that do not dephosphorylate glucose, G6P is converted into glycogen for storage -Glucokinase: special form of special kinase in the liver -Km[glucose] ~= 5 mm (~blood glucose conc.)

Final step of Gluconeogenesis:

-The generation of free glucose in the liver -Only occurs in the liver -Glucose 6 phosphate is transported into the lumen of the endoplasmic reticulum -Glucose 6 phosphatase catalyzes the formation of glucose from G6P -An integral membrane protein on the inner side of the endoplasmic reticulum -In tissues that do not dephosphorylate glucose, G6P is converted into glycogen for storage. -liver has a special form of hexokinase called glucokinase -Km(glucose) ~= 5 uM glucose

what determines the melting point of a fatty acid? (number of double bonds)

-The more double bonds in a fatty acid the lower the melting point. -e.g. EPA has 5 double bonds and a melting point of -54 C

Structure of Photosystem I (PSI)

-The psaA and psaB subunits are shown in red and blue respectively -The "special pair" (p700) absorbs hv -Trimer of protomers (each protomer has 2 distinct subunits)

Remember that this enzyme is not very specific. what confers specificity to targets?

-The shape of the catalytic site to the substrate it attempts to bind to

E3: Dihydrolipoyl dehydrogenase (part 2)

-The two adjacent Cys-SH on E3 allow for disulfide exchange between dihydrolipoamide and the enzyme -Essentially E3 exchanges S-S bonds with dihydrolipoamide -This forms a thiolate anion that needs to be oxidized to reform the S-S bond

How does bacteria establish a proton gradient?

-They establish it across their cytoplasmic membrane into the periplasm

E3: Dihydrolipoyl dehydrogenase (part 3)

-Thiolate anion forms a charge-transfer complex, then a covalent adduct with FAD -results hydride transfer to FAD making the FADH- anion -At this point the enzyme returns to original form (S-S bond reformed)

what is a multicyclic cascade, and what purpose does it have?

-Through multiple cycles 1 enzyme can degrade many substrates -Allows for the amplification of signals and divergence in pathways

Model of E. Coli F1F0 ATPase Cont.

-Top down view of C ring and subunit a -Another view of the intermembrane half channels

Ferrodoxin/NADP+ Reductase Diagram

-Transfers two electrons and a proton to NADP+ to form NADPH in the stroma - 2e- from PSI via ferrodoxin -FAD is a cofactor

Fatty Acid Overview

-Triacylglycerol (triglyceride): fat storage -Fatty acids have six times the energy content than hydrated glycogen per gram -Twice the energy produced per carbon (38 vs 17 kJ/g) -esterified with glycerol to form acyl glycerols, glycerides, or glycerophospholipids In a 150 lbs person: -420,000 kJ (11 kg) in reserve fat -Stored in cytoplasm of adipose cells Four main roles of fatty acids: -fuel storage -building blocks for phospholipids -Many proteins are modified by covalent attachment of fatty acid(s) -Fatty acid derivatives --> hormones and intracellular messengers

Place the steps regarding fat digestion and absorption in the correct order. The abbreviation TAG is used for triacylglycerol (triglyceride).

-Triacylglycerols enter stomach -Undigested fats enter the small intestine -The gallbladder secretes bile into the small intestine -Pancreatic lipase hydrolyzes TAGs into lipid droplets -Insoluble lipids, in micelles, are absorbed through the lining of the small intestine -TAGs in chylomicrons enter the lymph system -TAGs in chylomicrons enter the blood stream -Triacylglycerols absorbed into cells

Transport of Acetyl CoA from mitochondria into cytosol.

-Tricarboxylic acid carrier -Acetyl CoA + OAA --> citrate -citrate can then be transported into the cytosol -ATP citrate lyase: citrate --> acetyl CoA + OAA

why might ACC or FAS be good targets for cancer drugs?

-Tumors require a large amount of fatty acid synthesis to produce precursors for membrane synthesis -FAS inhibitors prevent the cell from synthesizing fatty acids ACC inhibitors prevent the cell from transporting ketone bodies and thus will not be able to transport energy out of the mitochondria

Describe how the PPP can generate glycolytic intermediates?

-Uses glucose 6 phosphate (usually produced via glycolysis) -Fructose 6 phosphate and glyceraldehyde 3 phosphate can also enter in 2 places (glycolysis intermediates) -ribulose 5 phosphate can be interconverted to many intermediates -occurs in cytoplasm of plants so intermingling of intermediates can occur -Not a top down pathway -The intermediates produced are determined by the needs of and the intermediates available to the cell -oxidative phase: -produces ribulose 5 phosphate (precursor to sugar in DNA/RNA) -non-oxidative phase -reactions are reversible so chance for intermediates to enter at each step

Calmodulin Structure

-Very common Ca2+ sensor -Sometimes works alone, but more often as a subunit of protein complexes -relieves auto-inhibition of phosphorylase kinase -Two similar globular domains connected by an alpha helix -When Ca2+ is bound to calmodulin it becomes more ordered -different binding properties occur -Hydrophobic residue is exposed -Can then bind to target protein -Ca2+ binding --> ordered ea segment --> Moves hydrophobic segment --> can now bind to other proteins -Once Ca2+ binds it relieves the auto-inhibition of phosphorylase kinase (makes it active)

Remember that Viagra's application to treat ED was discovered by 'accident' - it is a phosphodiesterase inhibitor.

-Viagra inhibits phosphodiesterase and thus cAMP cannot be degraded

You have discovered a new micro-organism whose F1F0 ATPase consists of a C-ring that has 15 subunits, as opposed to the 9 in the human complex, and 12 in bacteria. How many protons does this organism need to bring back in from the outside to synthesize one ATP? b. How many moles of ATP could it theoretically synthesize per mole of NADH, assuming the reaction is 100 % efficient?

-We need 15 H+ for one full turn -One turn makes 3 ATP -5 H+ per ATP

(3) Hormonal Control through Phosphorylation

-Whats regulates the regulator? "kinase kinase" Phosphorylase kinase (alpha, beta, gamma, delta)4: -gamma catalytic subunit (kinase) -Activated by Ca2+ (delta = calmodulin) -calmodulin: calcium binding module -Activated by phosphorylation (alpha, beta, regulatory subunits) -PKA: cAMP dependent protein kinase ("kinase kinase") -phosphorylates phosphorylate kinase -Makes active form -PP1: protein phosphatase 1 -dephosphorylates phosphorylase kinase -makes inactive form

cAMP Dependent Protein Kinase Activation

-When active gluconegenesis wins -Low blood glucose --> increased gluconeogenesis -Amplification: -1 glucagon --> many cAMP -Many PKA activated -PKA phosphorylates many targets SUMMARY: Glucogenesis is favored in a fasting state

Glycolysis and Gluconeogenesis Reciprocal Regulation Diagram

-When citrate is high energy is high -Citrate proportional to energy -Citrate forms in the TCA cycle

Control of the Synthesis and Degradation of Fructose 2,6 biphosphate

-When glucose is abundant fructose 2,6 biphosphate is high -when glucose is scarce fructose 2,6 biphosphate is low -The enzyme is regulated through phosphorylation and dephosphorylation

what is a substrate cycle, and why does it help when it is 'leaky'?

-When two metabolic pathways run simultaneously in opposite directions -e.g. the phosphorylation of F 6 P to F 1,6 bP and back to F 6 P -Sometimes called futile cycles -Neither metabolic pathway is fully active so one wins over slightly. (one leaks over the other) -These pathways enhance metabolic signals -A small change in the rates of the opposing reactions leads to a massive change in the net flux -A 20% in activation on both sides leads to a 380% change in net flux

G6P Dehydrogenase Deficiency

-X linked - ~400 million ppl -5-25% of the population in Africa, tropical Asia, Middle East, Mediterranean -Peroxides damage cell walls and Hb becomes oxidized -Jaundice: accumulation of biliverdin since not as much NADPH is produced -Protects against plasmodium falciparum (malaria)

Why does it make sense that Rubisco is regulated by both the pH and Mg ions? Where is Rubisco found in plants?

-[H+] decreases and [Mg] increase in the stroma when light reactions are taking place -It is then optimal to have rubisco activity -Found in the chloroplast -encoded by the chloroplast genome

Starch digestion:

-a-amylase --> di/trisaccharides --> a-glucosidasen --> debranching enzyme --> glucose

How are the levels of cAMP determined (activity of adenylate cyclase : phosphodiesterase)

-adenylate cyclase hydrolyzes (produces) cAMP -Phosphodiesterase degrades cAMP -?

General Information about the pentose phosphate pathway

-also called the hexose monophosphate shunt -Takes G6P and makes NADH and ribose S phosphate -NADH: has reducing power to make stuff -Ribose S-phosphate: DNA/RNA -Bypasses: PG1, PFK, and aldolase

Fate of pyruvate (anaerobic)

-anaerobic conditions --> fermentation pyruvate --> lactic acid (Homolactic fermentation) (us) Pyruvate --> ethanol (in microorganisms) Consumes: 2 NADH (per glucose) Produces: 2 ATP (Per glucose) No reductive power harvested Need to make ATP fast to regenerate NAD+

mechanism of phosphoglucomutase (how is the high-energy phosphate bond conserved?) Cont.

-another image

Decarboxylation by pyruvate decarboxylase (part 2)

-bottom half of the image shown right -Drunken monkey hypothesis: liking alcohol comes from our ancestors (and all nectar/fruit eating animals) consuming fermented fruit.

Hormonal Control through Phosphorylation Cont.

-cAMP dependent protein kinase -PKA -regulated by cAMP (talked about above) -cAMP dependent kinase is a "kinase kinase" -cAMP inhibited by Viagra

Phosphorylase kinase: how is it activated by cAMP dependent protein kinase, and by calcium (calmodulin)

-cAMP dependent protein kinase phosphorylates phosphorylase kinase making 2 catalytic subunits -this activates phosphorylase kinase -Calmodulin changes conformation around Ca2+ and opens the catalytic domain of phosphorylase kinase -This conformational change allows phosphorylase kinase to bind to substrates

Chemiosmotic Hypothesis (Peter Mitchell) and PMF

-chemiosmotic hypothesis: proton gradient provides free energy for ATP synthesis -PMF: free energy available from dissipation of H+ gradient -H+(matrix) --> H+(ims): chemical gradient of H+ -Electrical gradient: separation of charge when ion passes across the membrane -Takes energy to pump H+ into IMS (across the concentration gradient)

Outer Mitochondrial Membrane (OMM)

-contains porins -Free passage of molecules up to 10 kDa -Facilitated diffusion -Smooth outer-membrane of the mitochondrion

Complex I: NADH: ubiquinone (CoQ) oxidoreductase Cont. x4

-cytoplasm portion shown in the image on the right -Distances for e- to "jump" are large -Direction of e- drops down from more -dEo' to more +dEo' -Spatially organized in protein

LHC structure:

-delocalized e- -reduced pyrrole -LHC come in repeating chains -LHC II of pea plants: -7 Chlorophyll a -5 Chlorophyll b -2 carotenoids

Diagram of complexes in membrane

-diagram in image on the right

Starch

-dietary source of glucose -a(1,4) links -a(1->6) branch points

Cytochromes

-electron transferring proteins that contain a heme prosthetic group. -The heme iron cycles between Fe2+ and Fe3+ as it accepts or donates electrons

Light reactions:

-energy from light elevates e- to excited state

phosphoglycerate kinase (PGK) Cont.

-enough energy to drive ATP fermentation -Delta G" value of half reactions and overall reaction are in the picture

Glycogen in the liver

-exports glucose to other tissues -storage of glycogen and degrades it to glucose -Synthesis of glucose by gluconeogenesis -synthesis and storage/degreadtion of glycogen are 2 separate events -Fed state: produces glycogen -Fasting state: produces glucose from glycogen Cori cycle produces glucose from lactate

allosteric regulation

-fast -substrate accumulation -Product regulation: inhibition -Effectors: ATP, ADP/AMP, metabolites -energy status

Glycogen synthesis is primed by glycogenin - what does it do, and how? what are its substrates?

-glycogenin is a primer to form glycogen -all glycogen granules have glycogenin at center -Has intrinsic glucosyl transferase activity -Links C1 to Tyr194 -Glycogenin binds UDP-glucose (UDPG) which then binds glycogen synthase. Glycogen synthase then extends the primer cleaving off UDP Substrates: -UDPG

Glycogenin and Glycogen Synthesis Overview

-glycogenin: dimer -glycogenin's nonreducing enol catalyzes the addiction (1) Glucosyl transfer from UDPG to Tyr194 (2) Binds glycogen synthase (3) glycogenin extends primer -UDP-Glucose --> UDP (4) glycogen synthase extends glycogen (5) branching enzyme branches -glycogen synthase binds after step 3 -Every glycogen granule has glycogenin at center

Light Harvesting Center II (LHC II) from pea chloroplasts

-homotrimeric protein (3 identical monomers) -High density of cofactors - Nearly 40% of the proteins non-hydrogen atoms comprise its chlorophylls and carotenoids -Top view of LHC II in image on the right

Plant Photosystem (Complex I)

-hv hits photosystem II (chla reduced) -Energy of LHC drives photooxidation of chla (p680) -chla+ reduced by oxidation of H2O --> O2 - p680: chle(red) -(minus 1 e-)-> chl(ox) -(H2O --> O2)-> chl(red) -(Much clearer in the diagram on the right)

Where are Pyruvate carboxylase and pyruvate dehydrogenase located

-in the same compartment (mito) on the same substrate

In thylakoid membrane

-in thylakoid membrane light harvesting complexes "attenuate" (absorb most visible light) -Very high absorption coefficients

Fluoroacetate

-inhibits aconitase -deadly to livestock and found in many plants

lactose intolerance

-lactase mutation deficiencies

What is the Cori cycle, and what purpose does it have?

-lactate produced by muscle glycolysis is transported to the liver via the bloodstream where it is converted to glucose by gluconeogenesis. -Blood stream carries glucose back to the muscles where it is stored as glycogen -Mobilization of glucose from lactate -Produces useful energy source from waste byproduct

Photooxidation

-light driven electron transport -Drives: NADPH synthesis, proton gradient

Metabolic pathway regulation

-metabolic pathways are highly regulated to prevent accumulation of intermediates -

how is this pathway linked into isoprenoid synthesis (Mevalonate pathway)

-mevalonate pathway effects hydroxymethylglutaryl CoA synthase through isoprenoids

Human Pyruvate Kinase

-muscle at rest: glycolysis inhibited through high ATP/AMP ration inhibiting pyruvate kinase -During exercise: glycolysis stimulated through the cori cycle (glycogenesis) of fermentation

Photosynthesis in the dark:

-non-photosynthetic parts of plants

Pentose Phosphate Pathway Diagram

-oxidative phase -regenerative phase -Non-oxidative synthesis of Rib 5P, GAP, F6P -All regenerative phase reactions are at equilibrium

P680 Oxidation

-p680 lacks e- gets it back from water -2H2O --> O2 + 4e- + 4H+ (OEC) -P680 half reactions shown in the image on the right

Diagram of electron flow from H2O to NADP+ in photosynthesis

-p680*: strong oxidant -p700*: strong reducing agent -diagram has the shape of a Z

Photosystem I Charge Figure

-p700: chla dimer -Ao: A special chla -A1: phylloquinone (vitamine K) -Fx, FA, FB: Fe-S clusters -Fd: Ferrodoxin -Starts at the excitement of p700 and ends at NADP+ reductase (makes NADPH)

Cataplerotic Reactions

-pathways that use citric acid cycle intermediates -TCA cycle intermediates depleted for anabolic synthesis

mechanism of phosphoglucomutase (how is the high-energy phosphate bond conserved?)

-phosphate binds to glucose and then rebinds to the enzyme

Monod-Wyman-Changeux model of allostery

-phosphofructokinase 1 is the committed step -F6P + ATP --> F 1,6BP + ADP -PEP is tensed (inhibitor) end product -ADP is relaxed (activator) signals low energy - ~20 effectors -ATP is tense and AMP/ADP are relaxed -Citrate (TCA intermediate) is tense and F 2,6BP (especially made to regulate!) is relaxed

Succinyl CoA Synthetase: Cont.

-phosphorylated enzyme intermediate -Reaction mech. in diagram on the left

Complex III: Coenzyme Q-cytochrome C Oxidoreductase (Cytochrome bc1 complex) Cont. x3

-porphyrin ring has ~20 dislocalized e- -cytochrome proteins (isoprene) bind to heme prosthetic groups (no isoprene)

how are odd chain fatty acids synthesized (and remember how they are broken down, as well!)

-propionyl CoA is used as a primer in place of acetyl CoA

A few days after starting an extremely restrictive "no‑carb" fat‑based diet, an otherwise healthy man begins to feel tired and weak. You suggest that the man add some carbohydrates to his diet. Despite your explanation that "fats burn in the flame of carbohydrates," the man still refuses to consume carbohydrates. Consider other ways in which the man could supplement his diet to improve his metabolic health. Select all the compounds that could improve this man's ability to metabolize fats.

-pyruvate -glycerol -succinyl CoA

What are the reactions that allow the conversion of cytosolic NADH into NADPH during fatty acid biosynthesis? What enzymes are required? What is the sum of these reactions?

-pyruvate + CO2 + ATP + H2O ⟶ oxaloacetate + ADP + Pi + 2H+ -oxaloacetate + NADH + H+↽−−⇀ malate + NAD+ -malate + NADP+ ⟶ pyruvate + CO2 + NADPH -pyruvate carboxylase -malate dehydrogenase -malic enzyme -NADP+ + NADH + ATP + H2O ⟶ NADPH + NAD+ + ADP + Pi + H+

Stage 1 of TCA Cycle

-pyruvate --> Acetyl-CoA -pyruvate dehydrogenase complex (PDC) -High energy S-C bond: activated acetate carrier -3 enzymes: 3 enzyme bound substrates -NAD+ and Coenzyme A are substrate cofactors -E1: Pyruvate dehydrogenase: 24 subunits, TPP (cofact.) -E2: Dihydrolipoyl transacetylase: 24, lipoamide -E3: Dihydrolipoyl dehydrogenase: 12, FAD -E. Coli 60 subunits, 4600 KDa, 300 A in diameter (ribosomal) -Consists of 3 steps: 1) Decarboxylation 2) Oxidation 3) Transfer to CoA

alcoholic fermentation

-pyruvate --> acetaldehyde --> ethanol -pyruvate decarboxylase: Pyruvate --> acetaldehyde -Decarboxylates the a-keto group -alcohol dehydrogenase: acetaldehyde --> ethanol

What are the four regulatory points in the TCA cycle? How does energy levels in the cell affect the regulation?

-pyruvate dehydrogenase (PDH) complex -citrate synthase -isocitrate dehydrogenase -alphaketoglutarate dehydrogenase -When energy is high TCA cycle is inhibited -When energy is low citric acid cycle is activated

Cyclic Electron Transport

-red light -Driven only by hv --> PSI -Electrons from PSI cycle back to Cyt. b6-F complex and plastoquinone -Drives H+ transport, ATP synthesis -No NADPH synthesis (if NADPH/NADP+ ratio is high) -No O2 synthesis (like anoxygenic bacteria)

(2) Phosphoglucomutase

-remains bound on enzyme -similar to phosphoglycerate mutase 3PG <--> 2 PG -Both use hydrolysis -Would need ATP -Saves the high energy bond

energy balance of fatty acid oxidation (dont forget expenditure for initial activation of fatty acid) - how much ATP can we make, for example, from a C20 fatty acid? how would this look like if we started from a triglyceride?

-repetitions (n carbons/2 -1) -acetyl-SCoA (carbons/2) -1 NADH and 1 FADH2 per repetition - (I) repetitions(NADH*2.5) + repetitions(FADH2*1.5) - acetyl-SCoA(1 ATP + (2.5 ATP/NADH * 3NADH) + (1.5 ATP/FADH2 * 1 FADH2)) (II) acetyl-SCoA(10 ATP) -(I) + (II) -2 ATP (energy cost) = net ATP -example in the problem: -9 repetitions, 10 acetyl-SCoA -9(2.5) + 9(1.5) = 36 ATP -10(10) = 100 ATP - 36 + 100 -2 = 134 ATP for a C20 fatty acid -Rule of thumb: an extension of a fatty acid chain by 2 carbons allows it to make 14 more ATP -e.g. C16 = 104, C18 =120, C20 =134... -For a triglyceride: -multiply the number of ATP produced by each chain (like above) by 3 -add 15 ATP to account for pyruvate holding the chains together

Pyruvate kinase Cont.

-resonance stabilized Mech. in picture

similarities and differences between fatty acid beta oxidation and biosynthesis

-shown in image above -Synthesis partially shown in image

Alcohol Dehydrogenase (yeast: YADH)

-similar mech. to lactate dehydrogenase (mech. not shown in the picture) -LADH: liver alcohol dehydrogenase -detox reaction -acetaldehyde builds up causing asian flush -acetic acid (product) causes blood vessel dilation

Isoforms:

-slightly different versions of the same protein encoded by different genes -e.g. : liver glucokinase (lower affinity for glucose)

genetic regulation

-slow -Isoforms of enzymes: genetic disorders -Tfx. regulation?

ATP in synthesis of: Nucleic acids, proteins, polysaccharides

-tRNA charging? -ribosomes -DNA/RNA polymerization

Chemiosmotic Hypothesis

-the discovery that ATP synthesis is powered by a proton gradient is one of the two major biological advancements of the 20th (other is structure of DNA) -Theory was made by Peter Mitchell -Very eccentric and somewhat controversial -Made much of his research at a restored manor house where his research was funded in part by a herd of dairy cows

Alpha Beta Bond Cleavage Cont.

-thiolase ~= hydrolysis -Reverse claisen condensation in step one -Enolate anion is resonance stabilized in step 2 -tetrahedral intermediate in the last step -Fatty acyl CoA(n) + CoA + NAD+ + FAD + H2O --> Fatty acyl CoA(n-2) + acetyl CoA + NADH + FADH2

Decarboxylation by pyruvate decarboxylase (part 1)

-top half of the image shown right -pyruvate decarboxylase mech. shown in the two images -decarboxylation requires building up of negative charge on carbonyl in transition state

lactate dehydrogenase (LDH) (homolactic)

-used in humans -Tetramer of M-type (muscle) LDH-A: -Better adapted for pyruvate --> lactate -H-type (heart) LDH-B: -Better adapted for lactate --> pyruvate -regulation by isoforms of the enzyme (different needs in different tissues) -Mech. shown in picture -Glucose --> --> --> 2x lactate --> blood --> liver --> glucose

ATP has a high phosphoryl-transfer potential because of four key factors:

1) Pi stabilized by resonance 2) 4 negative charges (repulsion) 3) favorable entropy 4) Stabilization of products by hydration

6 Stages of Gluconeogenesis

1) Pyruvate --> PEP (bypasses pyruvate kinase) 2) PEP --> Fructose 1,6 biphosphate 3) Fructose 1,6 biphosphate --> Fructose 6-phosphate HPO42- (bypasses Phosphofructokinase) 4) Fructose 6-Phosphate --> Glucose 6-Phosphate 5) Glucose 6-Phosphate --> Glucose + Pi (bypasses hexokinase)

Four Different modes of operation of the Pentose Phosphate Pathway

1) Ribose 5 Phosphate needs exceed the need for NADPH 2) The NADPH and ribose 5 phosphate needs are balanced 3) More NADH is needed than ribose 5 phosphate 4)NADPH and ATP are both required -Modes 1 and 2 are utilized by cancer cells -PPP is required for rapidly dividing cells -Rapidly dividing cells need ribose 5 phosphate for nucleic acid synthesis and NADPH for fatty acid and membrane synthesis (e.g. cancer cells) -Glyolytic intermediates are diverted to the PPP by the expression of pyruvate kinase isozyme (PKM), which has low catalytic activity

Glycolysis stages:

1. Preparatory Stage: -"Energizing" - 6C glucose, phosphates added, split into 2 3C glucose phosphate, spending 2 ATP in process (substrate level phosphorylation). - 2GP are made (2 3C glucose phosphate) (2 GAP) -2ATP --> 2ADP 2. Energy Conserving Stage: - 2GP (2 GAP) are phosphorylated, another phosphate is added - 4 phosphate added to 4ADP to make 4ATP - End with 2 3C pyruvic acid and 4 ATP & 2NADH -4 ADP --> 4ATP -2NAD+ --> 2NADH SUM: 2 ATP, 2 NADH

How does H+ translocation drive ATP synthesis

1. Translocation of H+ from IMS ("out") to matrix ("in") 2. Catalysis of ADP + Pi --> ATP + H2O 3. Coupling of H+ translocation to ATP synthesis -Asymmetry of F1 a3b3 subunits -b subunit forms 3 different conformers -ATP bound (L) ("lose") -ATP bound (T) ("tense") -Empty (O) ("open")

What are the three steps of the calvin cycle?

1. carbon fixation 2. reduction 3. regeneration (of rubisco the CO2 acceptor)

Calculate the number of ATPs generated from one saturated 18-carbon fatty acid. For this question, assume that each NADH molecule generates 2.5 ATPs and that each FADH2 molecule generates 1.5 ATPs.

120 ATPs

Order the components of the ETC to outline the flow of electrons from NADH to O2 (put correct numbers).____Q-cytochrome c oxidoreductase ___ubiquinone ___cytochrome c ____NADH-Q oxidoreductase ___Cyt.c oxidase

1: NADH-Q oxidoreductase 2: ubiquinone 3: Q-cytochrome c oxidoreductase 4: cytochrome c 5: Cytochrome c oxidase

Krebs Cycle (Citric Acid Cycle): over all reaction

2 acetyl groups + 6NAD+ + 2FAD + 2ADP + 2Pi --> 4CO2 + 6NADH + 6H+ + 2FADH2 + 2ATP -generates: 8 NADH, 2 FADH2, 2 GTP (= ATP) per glucose

Select the results from having few or no a 1,6 linkages in glycogen: -Glycogen solubility would increase -Glycogen degradation would slow down -Glycogen synthesis would be faster -maintaining proper blood sugar levels would be more difficult

2 and 4 ( the alpha 1,6 linkages are the crosslinks that allow branching to occur) -Glycogen degradation would slow down -Maintaining proper blood sugar levels would be more difficult

Mode 4: Both NADPH and ATP are required

3G6P + 6NADP+ + 5NAD+ + 5Pi + 8ADP --> 5pyruvate + 3CO2 + 6NADPH + 5NADH + 8ATP + 2H2O + 8H+

Blood --> Cell Glucose transportation (Point of regulation)

4 main types of glucose transportation: (GLUT 1-4) -GLUT 1: High affinity transporter expressed in cell types with barrier functions. (insulin independent) -GLUT 2: High capacity low affinity transporter. used as glucose sensor in pancreas. used in liver and small intestine (insulin independent) -GLUT 3: High affinity transport used in the brain/CNS (insulin independent) -GLUT 4: High affinity transporter used in skeletal muscle, adipose tissue, and heart. Number of GLUT 4 transporters increase on the cell surface porportionally to glucose levels (insulin sensitive)

On average, how many glucose 1‑phosphate molecules will be released from a single glycogen branch at its nonreducing end before glycogen phosphorylase cannot cleave that branch any further?

6

Calculate the number of repetitions of the β‑oxidation pathway required to fully convert a 18-carbon activated fatty acid to acetyl‑SCoA molecules. Calculate the number of acetyl‑SCoA molecules generated by complete β oxidation of a 18-carbon activated fatty acid.

8 repetitions (n carbons/2 -1) 9 acetyl-SCoA (carbons/2)

ATP ~= GTP

ADP + GTP <--> ATP + GDP

oxidative phosphorylation

ATP Synthase: ADP + HPO4- --> ATP + H2O (Driving by H+ gradient)

TCA Overall Reaction

Acetyl CoA + 3NAD+ + FAD + ADP + Pi + 2H2O --> 2CO2 + 3NADH + FADH2 + ATP + 2H+ + CoA -Thioester cleavage powers ATP generation

what is a processive enzyme?

An enzyme that can catalyze subsequent reactions before releasing its substrate. -e.g. glycogen phosphorylase binds 4-5 units of glycogen and can phosphorylate several glucose units before it must rebind

NADP+/NADPH

Anabolism

Question in image on the right:

Answer below:

Four Modes of PPP:

Below

Recitation Worksheet Questions:

Below

Complex I:

Below:

Complexes I-IV of ETC

Below:

Light Reactions:

Below:

Mitochondrial Transport Proteins

Below:

Photosynthesis in the light:

Below:

Where is ATP used:

Below:

Cool Stuff on the Mitochondria

Below: (Subsection of ETC and oxidative phosphorylation)

Glycolysis (Mechanisms)

Below: -favorable since the last step is so favorable that it "pulls" the earlier unfavorable reactions -Conditions in the cell are usually more favorable than standard state conditions. -Why 10 steps?

Where does beta oxidation occur, which cellular compartment? Where does FA synthesis occur? Where are ketone bodies made, both body organ and cellular compartment?

Beta oxidation occurs in the mitochondria, FA synthesis happens in the cytosol. Ketone body synthesis happens in the mitochondria of the liver.

Understand the two enzymatic activities of glycogen debranching enzyme and why both are important.

Bifunctional -Two active sites: -a-1,4 glucanotransferase (GT) -shifts a block of 3 glucosyl residues from an outer branch to an inner one -Essentially takes the glucosyl residues in front of a branch point and moves them to an inner branch. think about cleaving little branches and adding them to the trunk of a tree until you get to the stump. -a-1,6 glucosidase (GC) -cleaves the glucose residue from the branch to form a linear branch and a glucose -Essentially removes the nubs of the branches so you have a linear branch

Branching and debranching enzyme - what do they do?

Branching enzyme: -Creates new non-reducing ends -Can count and measure glucosyl residues -Branches glycogen granules Debranching enzyme: -Debranches glycogen to free up glucose (last residue of the branch) and add glucosyl residues to main branch -This allows for further breakdown of glycogen -Like debranching a tree.

ETC and Oxidative Phosphorylation: Overall Reaction

C6H12O6 (glucose) --> --> --> 6Co2 + 24e- (stored in 10NADH and 2FADH2) 6Co2 + 24e- + 24H+ --> 12H2O -O2 consumed in mitochondria -How are the electrons from NADH and FADH2 transferred to O2? -How does this drive ATP synthesis?

CHO catabolism (respiration)

CHO --> CO2 + ATP + NADH + NADPH -glycolysis, oxphosin, pentose phosphate -Just like in humans

Glucagon secretion is stimulated when blood glucose concentration decreases. Select all of the carbohydrate pathways that glucagon stimulates in the liver.

Carbohydrate pathways that glucagon stimulates in the liver: -Glycogenolysis (glycogen breakdown) -Gluconeogenesis

Processes that release energy from glucose:

Carbon oxidation C-C bond cleavage *The more reduced a molecule is the more energy we can get out of it.

Exergonic

Chemical reaction that release energy (highly favorable)

Fate of pyruvate (aerobic)

Citric acid Cycle (TCA cycle): Stage 1: pyruvate --> AcetylCoA + CO2 Stage 2: AcetylCoA --> --> --> CO2 Produces: 8 NaDH, 2 FADH2, 2 GTP (per glucose) Glycolysis + Citric acid Cycle: Glucose --> 6 CO2 (Breathe out) Produces: 10 NADH, 2 FADH2, 4 ATP

Cori Cycle Cont.

Cori Cycle: -Mobilization of glucose from lactate produced in skeletal muscle via gluconeogenesis in the liver -Liver: -Synthesis/storage of glycogen -Degradation of glycogen to glucose -Synthesis of glucose by gluconeogenesis Skeletal Muscle: -Storage of Glucose as Glycogen -Glycogen --> g6P --> glycolysis -Gluconeogenesis via the cori cycle -Glucose --> lactate (in skeletal muscle) generates 2 ATP -Lactate --> glucose (in liver) consumes 6 ATP -Both are NADH neutral Glucose alanine cycle: -Muscle: pyruvate --> alanine -Liver: alanine --> pyruvate (urea excreted as a by product... urea cycle) -Cooperation between glycolysis and gluconeogensis during a sprint in the diagram on the left.

flux of carbon between skeletal muscle, heart muscle, and liver

Cori cycle: mobilization of glucose from lactate produced in skeletal muscle via glucogenesis in the liver. glucose alanine cycle: mentioned above flux of carbon in fed and fasting states is well indicated in the diagram

What are the differences between cyclic and non-cyclic electron transport in photosynthesis

Cyclic: -leads to the formation of ATP and NADPH -e- go from H2O --> PSII --> PSI --> --> -->NADPH Noncyclic: -Only some ATP is produced -e- go from PSII --> PSI and back again

Pathway of Gluconeogenesis

Diagram in image

ATP Hydrolysis

Drives unfavorable reactions forward

What are the products of each round of beta oxidation? What are the total products for the complete oxidation of palmitoyl CoA?

Each round of beta-oxidation yields 1 Acetyl-Coa, 1 FADH2, and 1 NADH. For complete oxidation of 16C FA we get 8 Acetyl-Coa, 7 FADH2, and 7 NADH

In exercising muscle, glycogen degradation supplies the muscle with glucose‑6‑phosphate. In order to stimulate muscle glycogen degradation, protein phosphatase 1 (PP1) must be inhibited. Four of the five events are involved in the inactivation of PP1 in exercising muscle.

Event not involved in the inactivation of PP1 in exercising muscle: -Insulin initiates a protein kinase cascade that utilizes glycogen synthase kinase. Events involved in the inactivation of PP1 in exercising muscle: -Epinephrine initiates a cAMPcAMP signal transduction cascade that utilizes protein kinase A. -Protein kinase A phosphorylates an inhibitor of PP1. -Protein kinase A phosphorylates GM in the GM-PP1 complex, resulting in its dissociation. -Phosphorylated PP1 inhibitor binds to PP1, facilitating glycogen degradation by phosphorylase a.

Regardless of its efficacy, what is the reasoning behind carbohydrate loading

Excess glucose is stored as muscle or liver glycogen, which can be broken down to supply energy during the event

What is the electron acceptor used for going from an alkane to an alkene?

FAD

Match each characteristic to the synthesis or oxidation of fatty acids.

Fatty acid synthesis: -occurs in the cytoplasm -Uses NADPH -Acyl carrier protein is the acyl carrier -involves the conversion of a carbonyl to a methylene -Enzymes are organized in a multienzyme complex Fatty acid oxidation: -Uses NAD+ -Uses FAD -Occurs in the mitochondria -Coenzyme A is the acyl carrier -Involves the conversion of a methylene to a carbonyl

Sucrose:

Fructose + glucose

What are the four main roles of fatty acids?

Fuel storage, building blocks for phospholipids, protein modifications, and to create fatty acid derivatives such as hormones and intracellular

Mode 3: Much more NADPH than ribose 5-phosphate is required

G6P + 12NADP+ + 7H2O --> 6CO2 + 12NADPH + 12H+ + Pi

Mode 2: The needs for NADPH and Ribose 5-phosphate are balanced

G6P + 2NADP+ + H2O --> ribose 5-phosphate + 2NADPH + 2H+ + CO2

Under conditions of low blood sugar, what hormone will predominantly be released? What does this cause to happen to beta oxidation and FA synthesis?

Glucagon is released. Increases beta oxidation, decreases FA synthesis

The hormone glucagon and the hormone and neurotransmitter epinephrine aid in maintaining blood glucose levels. Classify the statements as describing glucagon only, epinephrine only, or both glucagon and epinephrine.

Glucagon: -secreted by pancreatic a cells -polypeptide Epinephrine: -derived from tyrosine -stimulates muscle cells to break down glycogen -released in response Both: -released in response to a drop in blood glucose levels -inhibits glycolysis and stimulates gluconeogenesis in the liver -the receptor has seven transmembrane a helices

two fates of pyruvate in mitochondria, and how its fate is determined based on energy levels and needs of the cell

Gluconeogenesis: -Pyruvate is carboxylated to form oxaloacetate by pyruvate carboxylase -OAA is then reduced to malate and shuttled to the cytoplasm to produce phosphophenolpyruvate Glycolysis: -Pyruvate is converted to Acetyl CoA by PDH -Pyruvate is converted to OAA in the fed state and converted to acetyl CoA in a fasting state -PDH is suppressed when carbohydrate levels are low promoting gluconeogenesis

Overall: Glycolysic + TCAcycle + e- transport/oxidative phosphorylation

Glucose + 6 O2 --> 6 CO2 + 6 H2O => 32 ATP

Energetics of Glycogen Synthesis

Glucose + ATP --> G6P + ADP (-17 kJ/mol) G6P --> G1P (7 kJ/mol) G1P + UTP + H2O --> UDPG + 2Pi (-19 kJ/mol) Glycogen(n) + UDPG --> glycogen(n+1) + UDP (-13 kJ/mol) Overall: -Glucose + ATP + UTP + glycogen(n) + H2O --> glycogen(n+1) + ADP + UDP + 2Pi (-42 kJ/mol) -Favorable -addition of each glucose costs 2 ATP -UDP + ATP <--> UTP + ADP (dG0' ~ 0) -nucleoside diphosphatekinase is the enzyme -Sugar nucleotide intermediate

High fructose corn syrup

Glucose + fructose

'balance the reaction' - going from glucose to glycogen n+1 and from glycogen to glucose, n-1

Glucose --> glycogen(n+1): Glucose + ATP + UTP + glycogen(n) + H2O --> glycogen(n+1) + ADP + UDP + 2Pi (-42 kJ/mol) Glycogen(n) --> glycogen(n-1) + Glucose: Glycogen(n+1) + H2O --> glycogen(n) + glucose (-18 kJ/mol) Overall: ATP + UTP + 2H2O --> ADP + UDP + 2Pi (-24 kJ/mol?)

Malate/a-ketoglutarate carrier and glycerophosphate shuttle Cont.

Glycerophosphate Shuttle: -Brain/skeletal muscle -less efficient -Not a transporter -Irreversible: can go across concentration gradient -glycerophosphate shuttle shown in image on the right

Glycogen phosphorylase catalyzes a __________ reaction that breaks __________ in glycogen. Phosphorylase is one of the enzymes of glycogenolysis that directly generates __________ In the fasting state, the hormone glucagon ________ the enzyme, resulting in __________ in blood glucose levels.

Glycogen phosphorylase catalyzes a phosphorolysis reaction that breaks α‑1,4 linkages in glycogen. Phosphorylase is one of the enzymes of glycogenolysis that directly generates glucose 1‑phosphate. In the fasting state, the hormone glucagon stimulates the enzyme, resulting in an increase in blood glucose levels.

(3) Glycogen Synthase

Glycogen(n) + UDP-G --> glycogen(n+1) + UDP -Group transfer of glucose dG0' ~= -13 kJ/mol -Driven by the hydrolysis of the sugar nucleotide

oxidation in glycolysis vs. pyruvate breakdown

Glycolysis: not a lot of oxidation pyruvate --> TCA Cycle --> CO2 lots of oxidation in TCA

Regulation of Glycolysis:

Goals: - Balance supply + demand - homeostasis - respond to entropy needs -Flux control points: -delta G - If +delta G then change conc. of reactants or couple reaction to favorable reaction -Delta G of reactions in glycolysis in picture

Glycolytic enzymes that require ATP

Hexokinase, Phosphofructokinase

What is the driving force for ADP-glucose pyrophosphorylase reaction

Hydrolysis of pyrophosphate

Identify the characteristics of allosteric regulation of glycogen phosphorylase in the muscle and liver

In Muscle: -Activation generates glucose for the cell. -b form is the default state. -AMP binding induces R state. In Liver: -Activation liberates glucose for export. -a form is the default state. -Glucose binding induces T state.

Balance the equations for glycogen degradation

In image

Complete the reactions which show the transfer of glucose to a growing glycogen chain.

In image

Complete the sentences to correctly describe steps in fatty acid synthesis.

In image

During glycogenolysis, glycogen is broken down and converted to glucose 6‑phosphate, which can enter glycolysis or be used by the liver to raise blood glucose levels. Complete the sentences describing glycogen breakdown. Some terms will be used more than once, and two terms will not be used at all.

In image

Insulin receptor: dimerizes upon binding of insulin --> activates insulin stimulated protein kinases (ISPK), and many other insulin receptor substrates (IRS) through phosphorylation of tyrosine. Remember that insulin receptor is a tyrosine kinases

In image

Phosphorylation and Dephosphorylation Systems of Glycogen Metabolism in Muscle

In image

Table of Glycogen Storage Diseases

In image

The diagram shows the reactions of the β‑oxidation pathway. Label the reaction types on the diagram.

In image

Water is prevented from entering the active site of glycogen phosphorylase. What are the advantages of excluding water from the active site

In image

Glycogen synthase may be regulated by covalent modification and/or allosteric control. Label the diagram with the appropriate terms to describe glycogen synthase regulation. Note that some texts use glycogen synthase I instead of glycogen synthase a and glycogen synthase D instead of glycogen synthase b.

In the image on the right

glucose absorption:

In the small intestine via glucose transporters

Glycogen synthase catalyzes glycogen synthesis. Determine whether each example is associated with an increase or decrease in glycogen synthase activity.

Increased glycogen synthase activity -insulin -activation of phosphoprotein phosphatase (PP1) -phosphorylation of glycogen synthase kinase (GSK, or GSK3) Decreased glycogen synthase activity -phosphorylation of glycogen synthase -subunit dissociation of cAMP-dependent protein kinase (PKA) -phosphorylation (inactivation) of phosphoprotein phosphatase 1 (PP1) by PKA

When blood glucose is low, the pancreas releases glucagon, a peptide hormone which stimulates the liver to produce and excrete glucose. Binding of the hormone to its receptor triggers a "second messenger" cascade pathway that results in a change in the concentration or activity of many enzymes and signaling intermediates. The activity of some pathway enzymes is altered, as is the direction of net flow through some pathways. How does glucagon stimulation affect the concentration or activity of the given signaling intermediates and enzymes?

Increases: -adenyl cyclase -[3',5'-cyclic AMP] -protein kinase A -phosphorylase b kinase -glycogen synthase kinase -fructose-2,6-biphosphatase Decreases: -[fructose-2,6-biphosphate] -phosphofructokinase-2 Glucagon stimulation decreases which pathway enzymes? Select all that apply: -glycogen synthase -pyruvate kinase -phosphofructokinase-1 Glucagon stimulates which pathways? Select all that apply. -gluconeogenesis -glycogenolysis (glycogen breakdown)

Where does the ETC and oxidative phosphorylation occur?

Inner Mitochondrial membrane

Under conditions of high blood sugar, what hormone will predominantly be released? What does this cause to happen to beta oxidation and FA synthesis?

Insulin. Decreased beta oxidation, increased FA synthesis.

glucose transporters in epithelial cells

Into epithelial cells: -Symporter: SGLI glucose (or galactose) with Na+ -GLUT5: fructose Into blood from epithelial cells: -Na+/K+ ATPase -GLUT2: (concentration gradient) glucose, glacatose, fructose

What is the advantage of phosphorylation having opposite effects on glycogen synthesis and breakdown?

It prevents useless expenditure of energy

What would happen if G-protein hydrolyzed GTP at a fast rate? -It would get degraded rapidly -It would waste energy -It would not be able to stay bound to the GPCR -It would not be able to activate AC (adenylate cyclase)

It would not be able to activate AC (adenylate cyclase)

What is the purpose of ketone bodies? How are ketone bodies synthesized? How many Acetyl-CoA units does it require?

Ketone bodies provide an additional energy source when glucose levels are low, especially for tissues that can't metabolize triglycerides like the brain. It takes 3 Acetyl-Coa units to synthesize HMG-CoA.

fatty acid esterification and de-esterification by acyl glycerol transferases and lipases, phospholipases

Lipases: -Deesterification -removes acyl groups -tri --> di --> monoacylglycerol Transferases: -Esterification -Adds acyl groups -mono --> di --> tri

fundamental difference between liver, muscle, and brain in their glycogen metabolism / storage capacity

Liver: makes and mostly exports glycogen while storing some Muscle: Stores glycogen and glucose but mostly protein Brain: almost no energy stores (emergency only) consumes glucose and glycogen at a high rate

Diagrams of Glycogen Phosphorylase a/b ratios

Liver: phosphorylase b (dephosphorylated) is active Muscle: phosphorylase a (phosphorlated) is active -In both cases the relaxed state is active

Fates of glucose 6 P.

Liver: to glucose through glucose 6 phosphatase -glucose is then treleased in the blood for use in other tissues Muscle/brain: to pyruvate through glycolysis -pyruvate is then converted to lactate during muscle contraction Pentose Phosphate Pathway: ribose and NADPH

Energy source for the liver

Makes glucose from non-CHO source

Malate/a-ketoglutarate carrier and glycerophosphate shuttle

Malate-Aspartate shuttle: -transports e- across membrane (IMM) -Malate transportation is fully reversible and goes with the concentration gradient -malate dehydrogenase: swapping of utility groups between AA and ketoacid -H+ goes with aspartate into the cytosol depleting the H+ gradient -Malate-aspartate shuttle in image on the right -electrons of cytosolic of NADH are transported to mitochondrial NAD+

Epinephrine is released in response to stress, and is a fight‑or‑flight hormone. Select all metabolic pathways that epinephrine stimulates:

Metabolic Pathways that epinephrine stimulates: -Glycolysis in muscle -glycogen breakdown (glycogenolysis) in liver and muscle -Gluconeogenesis in the liver -Lipolysis in adipose tissue

G6P dehydrogenase deficiency Cont.

Most common human enzymopathy. Mutant enzyme breaks down quickly. Body responds to deficiency by producing more enzyme in new RBCs. When oxidative stress increases, anemia can occur.

How does the regulation of protein phosphatase activity differ in muscle and liver? Why is it different?

Muscle: the interaction of the phosphatase with the glycogen granule is enhanced by cAMP, but inactivated by camp-dependent protein kinase, PKA, (through phosphorylation of M protein). then, an inhibitor (also activated by protein kinase, binds and shuts down the phosphatase for good. In liver: inhibited by binding to the active glycogen phosphorylase. When glucose is abundant, that shifts to the T state, which no longer binds the particles. PP1 is then free to activate glycogen synthase by dephosphorylation, and inactivate the T-state (already pretty dead) of glycogen phosphorylase by removing the P now P'lase b.

Which of the following mutations in muscle glycogen phosphorylase could result in a condition similar to McArdle disease

Mutant glycogen phosphorylase is unable to bind to AMP

Electron Transport Chain:

NADH --> NAD+ + H+ + 2e- FADH2 --> FAD + 2H+ + 2e- exergonic Used to establish H+ gradient across mitochondrial membrane.

Source of reducing power for fatty acid synthesis

NADH and NADPH are the source of reducing power for fatty acid synthesis

What are the four ways that light energy can be transferred or dissipated? Name an example of each in the process of light energy being absorbed by LHCs. Draw a Jablonski diagram representing these forms of energy dissipation.

NEED TO COME BACK AND COMPLETE THIS -Fluorescence -Internal conversion -Excitation transfer -photo oxidation NEED JABLONSKI DIAGRAM AND NAME OF EACH PROCESS

Auxotroph

Needs to consume CHO

Is food that contains mono- and diacylglycerols really 'low fat', 'low calorie'?

No, Chemically, all these glycerides are esters of glycerol and fatty acids which are metabolized in exactly the same way. Monoglycerides and diglycerides, like normal fats, also have 9 Calories per gram. The FDA regulations require reporting fatty acids expressed as triglycerides. In a strict interpretation, monoglycerides and diglycerides are not considered "fat", and information about the saturation of their fatty acid components is omitted from the nutrition label.

why do we need glycogen / branched glycogen; know the concepts of 'reducing end' and 'nonreducing end'

Non-reducing ends: glycogen grows and degrades from those ends (C4) Reducing ends: these ends can oxidize and reduce other molecules (C1)

If skeletal muscle cells had the same mutation, what would be the effects on skeletal muscle cells?

Nothing. Skeletal muscle cells lack glucose-6-phosphatase.

Photorespiration (O2 --> CO2)

O2 --> CO2 -Alternate dark reaction pathway -Occurs at high [O2], low CO2, High T -dissipates energy

Arrange in proper order the events of the signal‑transduction cascade for glycogen degradation in muscle.

Order of signal transduction pathway for glycogen degradation in muscle: -Contraction begins -phosphorylase kinase is partly activated by binding Ca2+ -epinephrine is released and binds to muscle B-andrenergic receptors. -The simulatory Ga protein dissociates and activates adenylate cyclase -Intracellular cAMP levels increase, which activates protein kinase A -phosphorylase kinase is phosphorylated on its a and b subunits -Glycogen phosphorylase b is converted to glycogen phosphorylase a -phosphorylysis of glycogen yields glucose 1-phosphate

Know the involved organs / enzyme and remember a bit about the Coris.

Organs involved: -Liver: gluconeogenesis (lactate --> glucose) -Muscle: makes lactate through contraction, stores glucose as glycogen enzyme involved: alanine aminotransferase (ALT) -High ALT level in the blood signals liver damage -Abundant liver enzyme Coris: -Immigrated to the U.S. in 1922 -Gerty was the first U.S. woman to win nobel prize in science (1947: medicine) -Gerty worked with her husband Carl and directors of the institute did not want that.

Oxphosin

Oxidative phosphorylation

E1: Pyruvate Dehydrogenase

PDH Kinase: Trigonal bipyramidal intermediate -E1-Ser (active) --> E1-phospho-Ser (inactive) PDH Phosphatase: -E1-phospho-Ser (E1-pSer) (inactive) --> E1-Ser (active) -Both reactions favorable in the direction written

Pentose Phosphate Pathway vs Calvin Cycle

Pentose Phosphate Pathway: -Decarboxylate and oxidize C6 --> generate NADPH -Generates a series of 3, 4, 5, 6, 7 carbon sugars (ribose 5 Phosphate) Calvin Cycle: -Carboxylate (carbon fixation) C5, cleave, reduce to 2xC3 with NADPH --> sucrose, starch -Generates a series of 3, 4, 5, 6, 7 carbon sugars (ribose 5 phosphate, regenerate ribulose 1,5 bisphosphate) Shared between the two pathways: -transketolases -transaldolases

Glycogen phosphorylase a and b, T and R.

Phosphorylase a: -liver -(T): inactive -(R): active Phosphorylase b: -muscle -(T): inactive -(R): active -Phosphorylase a is phosphorylated and phosphorylase b is unphosphorylated

Allosteric regulation of glycogen phosphorylase - what is the structural basis (Km effect)

Phosphorylase b (unphosphorylated): -ATP, G6P, and caffeine favor phosphorylase b (T) (inactive) -AMP favors (R) (active) Phosphorylase a (phosphorylated): -glucose favors (T) (inactive) Allosteric effector site: -AMP: activator -ATP, G6P: inhibitors

Blood disorders caused by genetic disorders affecting Glycolysis

Pyruvate kinase deficiency: -anemia -Step 10 of glycolysis -High 2,3 BPG low O2 -Autosomal recessive (very common) Hexokinase deficiency: -Step 1 of glycolysis -severe anemia -Low 2,3 BPG high O2 -Autosomal recessive (very rare) -Hemolysis

P Esters are used for:

Regulation, activating metabolites, regulate metabolites

Citrate Synthase

STEP 1 Oxaloacetate to Citrate

citrate synthase

STEP 1 acetyl CoA to citrate Start of first stage

pyruvate kinase

STEP 10: phosphoenolpyruvate to pyruvate -Last step of stage 2 and overall glycolysis -Named for the reverse reaction -Net production: 2 ATP, 2 NADH, 2 Pyruvate

Hexokinase

STEP 1: The enzymes that catalyzes the phosphorylation of glucose to form glucose-6-phosphate in the first step of glycolysis. This is one of the ain regulatory steps of this pathway. Hexokinase is feedback-inhibited by glucose-6-P. -G6P (glucose-6-P) gets gamma phosphate from ATP -Metal (Mg2+) assisted phosphoryl transfer -1) Base assisted catalysis: -OH more nucleophilic -2) Mg2+ assisted Phosphate transfer: gamma phosphate more electrophilic

Aconitase

STEP 2 Citrate to isocitrate -Dehydrogenation and rehydrogenation -2' alcohol to 3' alcohol - (2) -CH2-COO- is not the same as (4) -CH2-COO- -Went to decarboxylate... want keto group -3' alcohol cannot be oxidized so we need 2' alcohol -

Aconitase

STEP 2 citrate to isocitrate

phosphoglucose isomerase (PGI)

STEP 2: glucose-6-phosphate to fructose-6-phosphate -Why isomerization: -C6 --> 2C3 each C3 needs a P so we need a primary alcohol -Keto group is needed to cleave C-C bond

Isocitrate dehydrogenase

STEP 3 Isocitrate to alpha-ketoglutarate -easy and almost spontaneous (Delta Go' = -21 kj/mol) -carbanion intermediate -Glut = C5 -Oxalocitrate remains enzyme bound

isocitrate dehydrogenase

STEP 3 Isocitrate to aplha-ketoglutarate :rate limiting enzyme in krebs cycle

Phosphofructokinase

STEP 3: The enzyme that catalyzes the phosphorylation of fructose-6-phosphate to form fructose-1-6-bisphosphate in the third step of glycolysis. This is the main regulatory step of glycolysis. PFK is feedback-inhibited by ATP. -"committed step" (basically irreversible) -Quintessential allosterically regulated enzyme - E. Coli version is a tetramer of 4 identical subunits

Aplha-ketoglutarate dehydrogenase

STEP 4 alpha-ketoglutarate to succinyl CoA -Harder due to alpha keto group -"pass" energy from thioester to GTP

alphaketoglutarate dehydrogenase

STEP 4 alphaketoglutarate to succinyl CoA

Aldolase

STEP 4: cleaves fructose 1,6-bisphosphate into GAP and DHAP -aldol cleavage (aldol condensation occurs by the reverse mech.) C6 --> 2x C3

Succinyl CoA Synthetase:

STEP 5 Succinyl CoA to Succinate -"thiokinase" -One and only place for ATP generation in the TCA cycle. -substrate level phosphorylation

Succinyl CoA synthetase

STEP 5 succinyl-CoA to succinate

Triose phosphate isomerase (TIM)

STEP 5: -End of stage 1 -makes glyceraldehyde 3-phosphate (GAP/G3P) -High energy phosphate group -SUM of Stage 1: 2 Glucose --> 2x GAP 2 ATP consumed

Succinate Dehydrogenase

STEP 6 Succinate to Fumarate -hydride transfer -Succinate dehydrogenase has 3 Fe-S centers -Functions to transfer electrons to Ox Phosin -FAD is used here since free energy change is not sufficient to reduce NAD+

succinate dehydrogenase

STEP 6 succinate to fumarate

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH)

STEP 6: produces NADH which can feed into the ETC -Start of stage 2 -Aldehydic carbon goes from +1 --> +3

Fumerase

STEP 7 fumarate to malate -2' alcohol is added -No redox -Resonance stabilized -Rxn is driven forward by low [OA]

phosphoglycerate kinase (PGK)

STEP 7: 1,3-bisphosphoglycerate to 3-phosphoglycerate -substrate level phosphorylation

Fumarase

STEP 8 fumarate to malate

Malate Dehydrogenase

STEP 8 malate to oxaloacetate -Delta Go' = + 30 kj/mol -

phosphoglycerate mutase

STEP 8: 3-phosphoglycerate to 2-phosphoglycerate -2,3 BPG intermediate

malate dehydrogenase

STEP 9 malate to oxaloacetate End of 2nd stage

Enolase

STEP 9: 2-phosphoglycerate to phosphoenolpyruvate (PEP) -PEP has e- repulsion and, thus, super high entropy between the phosphate and carboxylic acid. -Overall delta G': -62 KJ (enough to make ADP --> ATP)

general acid-base catalysis

Side chain of AA (LYS, HIS) assists in proton transfer

If you were to design a low calorie fat substitute, what key features would you want? Hint: Why is sucrose-octaester (olestra) a low-calory fat substitute?

Similar features to fatty acids but make it indigestible

Km

Substrate concentration at 1/2 Vmax

Anabolism

Synthesis of building blocks using: ATP, NADPH (mainly)

Energetics of glycogen synthesis and breakdown

Synthesis: - -14 kJ/mol -favorable Breakdown: - -18 kJ/mol -favorable -Energetics of partial reactions are shown in the image

the direction of glycogen synthesis and breakdown.

Synthesis: fed state -glycogen to G1P Breakdown: fasting state -Activates glucose by forming a high energy sugar nucleotide -Glucose to branched glycogen

We have encountered reactions similar to the sequential oxidation, hydration, and oxidation reactions of fatty acid degradation earlier in our study of biochemistry. What other pathway employs this set of reactions?

TCA Cycle

Why is water excluded from the active site of glycogen phosphorylase, and how do you think this could be achieved?

The active site closes in order to prevent hydrolysis resulting in glucose rather than G6P

produced inhibition

The inhibition of an enzyme by a product that supersedes the enzyme in a metabolic pathway

feed-forward activation

The stimulation of an enzyme by an intermediate that precedes the enzyme in a metabolic pathway

Example of alternate reaction without thioester intermediate

Thioester intermediate coupling the reactions greatly reduces the activation energy of acyl phosphate formation. (1,3-BPG)

Andersen's disease is a deficiency of the glycogen branching enzyme (amylo(1,4 1,6) transglycosylase). This is one of the most severe glycogen storage diseases, resulting in death before age of 2 from progressive cirrhosis of the liver. Explain what the glycogen would look like, and speculate why this results in progressive liver damage.

This would cause the glycogen to be super long chains without any branching. It is recognized by the immune defense system and therefore results in liver damage.

How does ETC and oxidative phosphorylation occur?

Through the oxidation of NADH --> Membrane potential --> ATP synthesis

how is the first oxidative step integrated in the electron transport chain?

Through the use of NADH and FADH2 to form ATP

In glycogen metabolism, which molecule's synthesis reaction is by the hydrolysis of pyrophosphate

UDP-glucose

Red blood cells and Nerve Cells

Use only glucose

You have identified a patient whose heterotrimeric G protein has a 10-fold higher turnover rate in its GTPase activity. Its interaction with hormone-bound receptor is normal. Explain the downstream effect.

Very little cAMP would be made because the adenylate cyclase wouldn't remain active for very long. There would be less activation of glycogen breakdown as a result.

ATP mechanical work

Via actomyosin -Contractility: (contraction) -Cell Migration -Transport of stuff in cells (walking kinesin)

ATP in active transport

Via: Na+/K+ ATPase and Ca2+ ATPase -Neurotransmission -Nutrient Uptake -Cell to Cell communication

Debranching enzyme allows for the complete degradation of glycogen by glycogen phosphorylase. In eukaryotes, debranching enzyme is a bifunctional enzyme containing transferase and α‑1,6‑glucosidase activity. Transferase transfers three glucosyl residues from a limit dextrin to another part of the glycogen molecule. α‑1,6‑Glucosidase hydrolyzes the α‑1,6 bond that links the single glucosyl residue remaining at the branch point after the limit dextrin has been transferred. What is the product of α‑1,6‑glucosidase activity?

What is the product of α‑1,6‑glucosidase activity? -Glucose

In a skeletal muscle cell without any mutations, is any glucose generated from the breakdown of glycogen? If yes, does it accumulate?

Yes, skeletal muscle cells also have the glucosidase activity of glycogen debranching enzyme, but since hexokinase has such a low Km, they readily grab the glucose and send it down the glycolytic pathway.

In the liver, if we had a mutation that completely blocked glucose-6-phosphatase activity Would any glucose be generated from glycogen degradation?

Yes, the glucosidase activity of glycogen debranching enzyme hydrolyzes off the last glucose residue instead of phosphorylating off the last residue and according to the book, about 10% of the glucose residues are hydrolyzed off

Active transport of H+

a) (Q cycle) e- transport via plastoquinone (8H+/4e-) b) (OEC) 2H2O --> O2 + 4H+ (4H+/4e-) -total of 12H+/4e- -Protonmotive force (dP): Driving force for ATP synthesis -Free energy required to transfer H+ from the stroma to the lumen -Chemical gradient -Electrical gradient -pH(lumen) < pH(stroma) -dp = pH(lumen) - pH(stroma) < 0 (3-4 pH units) -(in mitochondria 0.7) -Equation for dG and values in diagram on the right -Protonmotive force: free energy available from dissipation of proton gradient -dpsi ~= 100 mV -(170 mV in mitochondria) -Counter flow of Mg2+ out of the lumen dissipates psi -Artificially high [H+] in lumen

Photooxidation

a) 2H2O --> O2 + 4e- + 4H+ (E0' = -0.82V) b) 2H+ + 4e- + 2NADP+ --> 2NADPH (E0' = -0.32V) -Overall: 2H2O + 2NADP+ --> O2 +2NADPH + 2H+ (EO' = -1.1 V) -dG0' = -nFdE0' -F = faraday constant 96 kJ/ V*mol -n = number of electrons -dG0' = 440 kJ/mol (for the reaction above...needs to be powered by sunlight)

structures of alpha-D and beta-D glucose, and the two types of linkages in glycogen

alpha D: -"away" -The C1 OH is down beta D: -"Both" -The C1 OH is up (same plane as CH2OH)

Answer the question using only your knowledge of glycogen metabolism enzymes. An infant is brought to a physician after having seizures. The infant had been noted as being very small for her age (below the 5th percentile for weight). During examination, the physician noted an enlarged abdomen (due to an enlarged liver). Tests show fasting hypoglycemia (low blood glucose concentration) and increased serum lipid levels. Liver and muscle biopsy revealed an increased amount of glycogen with short branches.

amylo-α-1,6-glucosidase (debranching enzyme)

ATP in CHO, lipid, AA, nucleotide synthesis

anabolism

Catabolism

break down of building blocks to generate: ATP, NADH, NADPH, FADH2

ATP in CHO, lipid, AA catabolism

breakdown

Which of these is a second messenger? -Epinephrine -G-protein -cAMP -Adenylate cyclase

cAMP

Warburg effect:

cancer cells preferentially use aerobic glycolysis while decreasing oxidative phosphorylation -Tumor uses lactate as the acidity promotes invasion and inhibits immune response -G6P used to NADPH through the pentose phosphate pathway -HIF: hypoxia inducing factor -transcription factor induced by hypoxia and anaerobic exercise -Mech. of fermentation shown in picture -table in the left of the picture shows proteins in glucose metabolism regulated by HIF -Table in the right of the picture shows starting and ending point of various fermentations

NAD+/NADH

catabolism

Hydrolases

catalyze cleavage with the addition of water

Oxidoreductases

catalyze oxidation-reduction reactions that involve the transfer of electrons (glyceraldehyde-3-phosphate dehydrogenase)

Lysases

catalyzes addition of groups to double bonds, or formation of double bonds by removal of groups. (aldolase, enolase)

Ligases

catalyzes condensation reactions with the input of ATP (forms C-C, C-S, C-O, C-N)

Isomerase

catalyzes the rearrangement of bonds within a single molecule (phosphohexose isomerase, triosephosphate isomerase, phosphoglycerate mutase)

CHO

cholesterol

pyruvate dehydrogenase (Mechanism)

converts pyruvate to acetyl-CoA. - stimulated by insulin - inhibited by acetyl-CoA

ATP hydolysis

energy from ATP hydrolysis is used to lower the entropy of the cell

GLUT 5

fructose transporter in the intestinal epithelium and spermatozoa

Irreversible reactions in glycolysis

glucokinase/hexokinase, Phosphofructokinase-1, pyruvate kinase

Suppose a researcher introduces a mutation into the glucosidase domain of the mammalian glycogen debranching enzyme. The mutation inhibits the activity of the glucosidase but does not affect the other functions of the enzyme. The researcher then introduces the mutated enzyme into mammalian cells that do not express wild type glycogen debranching enzyme. Predict the effect of the mutation on glycogen metabolism

glycogen molecules with branches containing a single glucose residue

Energetics of Glycogen Breakdown

glycogen(n+1) + Pi + --> glycogen(n) + G1P (dG0' ~= 3 kJ/mol) G1P --> G6P [in liver] (-7 kJ/mol) G6P + H2O --> glucose + Pi (-14 kJ/mol) Overall: -glycogen(n+1) + H2O --> glycogen(n) + glucose (-18 kJ/mol) -Favorable

allosteric enzymes

have both an active site for substrate binding and an allosteric site for binding of an allosteric effector (activator, inhibitor). -Very idealized representation of protein structure

Metabolic Map link

http://savi-cdn.saplinglearning.com/metabolicmap/index.html

Glycogen breakdown in image

in image

Table of Gluconeogensis Reactions

in image -G6P --> glucose (only in liver)

How would the following changes affect the rates of glycogen synthesis and glycogen breakdown? Write down which enzymes are regulated, and how.

increased cytosolic Ca2+: -Calcium activates calmodulin subunit in phosphorylase kinase and partially activates the enzyme. Phosphorylase kinase then phosphorylates phosphorylase to activate it. Phosphorylase kinase also phosphorylates glycogen synthase to inactivate it. This essentially turns on glycogen breakdown and inhibits glycogen synthesis. increased plasma insulin: -Insulin binds the insulin receptor which allows the phosphorylation of a number of target proteins including: phosphodiesterase and protein phosphatase 1. Activated phosphodiesterase hydrolyzes cAMP. As a result, cAMP can no longer activate PKA and glycogen breakdown is inhibited. When protein phosphatase 1 is phosphorylated, it dephosphorylates glycogen synthase which activates it and phosphorylates phosphorylase to inactivate it. Basically turns glycogen synthesis on and glycogen breakdown off.

Regulation:

metabolic pathways must be regulated to create homeostasis (stable biochemical environment)

Mutases

rearrangment of functional groups

NADH/NADPH

redox currency

Why the ETC and oxidative phosphorylation?

to produce ATP

Transferases

transfer functional groups from one substrate to another. (Require ATP... kinases) (Hexokinase, phosphofructokinase-1, phosphoglycerate kinase, pyruvate kinase)

Coenzyme Q

ubiquinone