Biochem Exam 3

thioesters

*high energy molecules because of inclusion of sulfur vs. general R-group Sulphur atom replaces oxygen Hydrolysis generates a carboxylic acid - Product is resonance stabilized - deltaG' is negative

anaerobic glycolysis: fermentation

- Generation of energy (ATP) without consuming oxygen or NAD+ - final reaction reduce pyruvate to another product - NADH is oxidized to make NAD+ so glycolysis can continue to run - No net change in oxidation state of the sugars - Reduction of pyruvate to another product - Regenerates NAD+ for further glycolysis under anaerobic conditions - The process is used in the production of food from beer to yogurt to soy sauce ( lactic acid fermentation and alcoholic fermentation)

Cryoelectronmicroscopy of PDC

- Samples are in a near-native frozen hydrated state - Low temperature protects biological specimens against radiation damage - Electrons have a smaller de Broglie wavelength and produce much higher resolution images than light The pyruvate dehydrogenase complex. (a) Cryoelectron micrograph of PDH complexes isolated from bovine kidney. In cryoelectron microscopy, biological samples are viewed at extremely low temperatures; this avoids potential artifacts introduced by the usual process of dehydrating, fixing, and staining.

glycerol 3-phosphate shuttle

- electrons are transferred from NADH to dihydroxyacetone phosphate (DHAP), forming glycerol 3-phosphate - these electrons can then be transferred to mitochondrial FAD, forming FADH2 brown fat & skeletal muscle, glucose → 30 net ATP (↓ efficiency means ↑ heat production) DHAP reduced to glycerol-3P to oxidize NADH in cytosol Then glycerol-3P re-oxidized to reduce FAD at mitochondrial inner membrane Q is reduced with electrons from FADH2 and QH2 goes to complex III Functions in muscle and brain

malate-aspartate shuttle

- electrons are transferred from NADH to oxaloacetate, forming malate - malate can then cross the inner mitochondrial membrane and transfer electrons to the mitochondrial NAD+, forming NADH heart & liver, most of the body, glucose → 32 net ATP functions in liver, kidney, heart

Pentose Phosphate Pathway

- utilizes Glucose-6P - the main products are NADPH and ribose 5-phsophate - NADPH is an electron donor > reductive biosynthesis of fatty acids and steroids > needed in liver , adipose , and erythrocytes - ribose-5-phosphate is a biosynthetic precursor of nucleotides > used in DNA and RNA synthesis > or synthesis of some coenzymes > needed in rapidly growing cells like bone marrow, skin and intestinal epithelium - pathway is in cytosol

Pyruvate to Phosphoenolpyruvate ( PEP )

1st bypass reaction - but takes two reactions to make phosphoenolpyruvate requires two energy-consuming steps deltaG' = 0.9 kJ/mol for both reactions FIRST step, pyruvate carboxylase converts pyruvate to oxaloacetate - carboxylation of pyruvate - requires bicarbonate and ATP > ATP used to phosphorylation bicarbonate to activate it - in eukaryotes, requires transport of pyruvate into the mitochondria SECOND step, phosphoenolpyruvate carboxykinase converts oxaloacetate to phosphoenolpyruvate - carboxyl group removed - phosphorylation using phosphate from GTP - loss of carboxyl and hydrolysis of phosphate from GTP both needed to produce the high energy compound - occurs in mitochondria or cytosol depending on the organism Pyruvate is first transported from the cytosol into mitochondria or is generated from alanine within mitochondria by transamination, in which the alpha amino group is transferred from alanine (leaving pyruvate) to an alpha keto carboxylic acid Then pyruvate carboxylase, a mitochondrial enzyme, converts the pyruvate to OXALOACETATE In the cytosol, oxaloacetate is converted to phosphoenolpyruvate by PEP carboxykinase. The CO2 incorporated in the pyruvate carboxylase reaction is lost here as CO2. This Mg2+ -dependent reaction requires Guanosine Triphosphate (GTP) as the phosphoryl group donor

Summarize glycolysis: What is used (total, not net)?

2 ATP, 1 glucose, 2 NAD+ 4 ADP and 2 phosphates are used

How many cycles of the citric acid cycle occur for every glucose molecule that goes through glycolysis?

2 cycles occur for every glucose molecule

How many protons are pumped by Complex IV per H2O produced?

2 protons pumped per H2O

gluconeogenesis is expensive

2 pyruvate + 4 ATP + 2 GTP + 2 NADH + 2 H+ + 4 H2O > glucose + 4 ADP + 2 GDP + 6 Pi + 2 NAD+ costs 4 ATP, 2 GTP, and 2 NADH physiologically NECESSARY brain, nervous system, and red blood cells generate ATP almost entirely from glucose when glycogen stores are depleted glucose must come from somewhere - during starvation or vigorous exercise plants use gluconeogenesis to make carbohydrates after carbon fixation

Summarize glycolysis: What is made (total, not net)?

2 pyruvate, 4 ATP, and 2 NADH are made in glycolysis.

simplified reactions combining 2 cycles of non-oxidative phase

2 ribose 5-P + 4 Xylulose 5-P > 4 Fructose 6-P + 2 Glyceraldehyde 3-P > 5 Glucose 6-P

How many protons are moved to the intermembrane space during the Q cycle? (Remember protons are not pumped at Complex III. They just leave the active site on the correct side of the membrane)

4 protons are moved to the IMS (2 are moved for each ubiquinol that is oxidized)

there are multiple glucose transporters

A Na+-glucose symporter and a glucose uniporter operate on opposite sides of intestinal epithelial cells Na+K+ ATPase uses ATP to pump ions against their concentration gradients Glucose transport in intestinal epithelial cells. Glucose is cotransported with Na+ across the apical plasma membrane into the epithelial cell. It moves through the cell to the basal surface, where it passes into the blood via GLUT2, a passive glucose uniporter. The Na+K+ ATPase continues to pump Na+ outward to maintain the Na+ gradient that drives glucose uptake.

pyruvate dehydrogenase complex ( PDC)

A group of three enzymes that decarboxylates pyruvate, creating an acetyl group and carbon dioxide. The acetyl group is then attached to coenzyme A to produce acetyl-CoA, a substrate in the Krebs cycle. In the process, NAD+ is reduced to NADH. The pyruvate dehydrogenase complex is the second stage of cellular respiration. PDC is a large (up to 10 MDa) multienzyme complex - pyruvate dehydrogenase ( E1) - dihydrolipoyl transacetylase ( E2) - dihydrolipoyl dehydrogenase ( E3) advantages of multienzyme complexes: - short distance between catalytic sites allows channeling of substrates from one catalytic site to another - channeling minimizes side reactions - regulation of activity of one subunit affects the entire complex

triacylglycerol

A lipid consisting of three fatty acids linked to one glycerol molecule; also called a fat or triglyceride. Glycerol and a triacylglycerol. The mixed triacylglycerol shown here has three different fatty acids attached to the glycerol backbone. When glycerol has different fatty acids at C-1 and C-3, C-2 is a chiral center (p. 17).

Name the two membrane phases. What types of fatty acids promote each one?

A membrane can be found in either the gel phase (liquid-ordered) or the fluid phase (liquid-disordered). A higher number of short, unsaturated fatty acids promote the fluid phase, whereas more long, saturated fatty acids promote the gel phase.

chemiosmotic theory

A model to explain the synthesis of ATP. The theory proposes that the energy for ATP synthesis originates from the electrochemical gradient of protons across a membrane The energy released by electron transport is used to transport protons against the electrochemical gradient Energy needed to phosphorylate ADP is provided by the flow of protons down the electrochemical gradient A different way to make ATP, not the result of a direct reaction between ADP and some high-energy phosphate carrier ADP + Pi -> ATP is Highly Thermodynamically Unfavorable The chemiosmotic mechanism for ATP synthesis. (a) In mitochondria, electrons move through a chain of membrane-bound carriers (the respiratory chain) spontaneously, driven by the high reduction potential of oxygen and the relatively low reduction potentials of the various reduced substrates (fuels) that undergo oxidation in the mitochondrion. Electron flow creates an electrochemical potential by the transmembrane movement of protons and positive charge. This electrochemical potential drives ATP synthesis by a membrane-bound enzyme, ATP synthase, that is fundamentally similar in both mitochondria and chloroplasts, and in bacteria and archaea as well.

multiple complexes associate together to form a respirasome

A putative respirasome composed of Complexes III and IV. (a) Purified supercomplexes containing Complexes III and IV, from yeast, visualized by electron microscopy after staining with uranyl acetate. The electron densities of hundreds of images were averaged to yield this composite view. (b) The x-ray-derived structures of one molecule of Complex III (red; from yeast) and two of Complex IV (green; from bovine heart) could be fitted to the electron-density map to suggest one possible mode of interaction of these complexes in a respirasome. This view is in the plane of the bilayer (yellow).

What is the number and direction of transport of molecules in a uniport, a symport and an antiport?

A uniport moves on molecule in one direction. A symport moves two or more molecules in the same direction. An antiport moves two or more molecules in opposite directions

ABC transporters use ATP hydrolysis to drive transport of substrates

ABC-ATP binding cassette ATP hydrolysis changes protein conformation and allows transport up a concentration gradient Two ABC transporters. (a) The multidrug transporter of animal cells (MDR1, also called P glycoprotein; PDB ID 3G60), responsible for pumping a variety of antitumor drugs out of human cells, has two homologous halves (blue and light blue), each with six transmembrane helices in two transmembrane domains (TMDs; blue), and a cytoplasmic nucleotide-binding domain (NBD; red). (b) The vitamin B12 importer BtuCD (PDB ID 1L7V) of E. coli is a homodimer with 10 transmembrane helices (blue, light blue) in each monomer and two NBDs (red) that extend into the cytoplasm. (c) Mechanisms proposed for the E. coli vitamin B12 ABC transporter coupling of ATP hydrolysis to transport. Substrate is brought to the transporter on the periplasmic side by a substrate-specific binding protein. With ATP bound to the NBD sites, the transporter is open to the outside (periplasm), but on substrate binding and ATP hydrolysis to ADP, a conformational change exposes the substrate to the inside surface, and it diffuses away from the transporter and into the cytosol.

What are the 3 molecules that are attached to a long chain fatty acid before it is transported into the mitochondrial matrix?

AMP, CoA-SH and carnitine are each attached to a long chain fatty acid at some point before it is transported into the mitochondrial matrix.

Electron Transport is coupled to ATP synthesis. What does this mean? Why does the inhibition of ATP synthase prevent electron transport from running?

ATP synthesis uses the proton motive force that set up by the electron transport, thus ATP synthesis requrires electron transport, and at the same time electron transport requred ATP synthesis in order to maintaine the proton gradient. Also, if electron transport is blocking it blocks the ATP synthesis. When ATP synthase is inhibited the proton gradient is rises and when it reaches the maximum concentration in the intermembrane space on the P side, it can not accept any more protons, and a lot a of energy needs to keep adding the protons to the P side, which become very thermodynamically unfavorable. Electron transport stops when protons can no longer be moved to the intermembrane space.

oxidation of fatty acids is a major energy source in many organisms

About one-third of our energy needs comes from dietary triacylglycerols About 80% of energy needs of mammalian heart and liver are met by oxidation of fatty acids Many hibernating animals, such as grizzly bears, rely almost exclusively on fats as their source of energy Plants use fats in seed germination

What is the final product of each cycle β-oxidation beta-oxidation that is used in citric acid cycle?

Acetyl CoA is the final product.

net result of the citric acid cycle

Acetyl-CoA + 3NAD+ + FAD + GDP + Pi + 2 H2O -> 2CO2 + 3NADH + FADH2 + GTP + CoA + 3H+ net oxidation of 2 carbons to CO2 - equivalent to 2 carbons of acetyl-CoA - but NOT the exact same carbons energy captured by electron transfer to NADH and FADH2 generates 1 GTP, which can be converted to ATP

net result of the citric acid cycle

Acetyl-CoA + 3NAD+ + FAD + GDP + Pi + 2 H2O -> 2CO2+ 3NADH + FADH2 + GTP + CoA + 3H+ Net oxidation of two carbons to CO2 - Equivalent to two carbons of acetyl-CoA (but NOT the exact same carbons) Energy captured by electron transfer to make NADH and FADH2 Generates 1 GTP, which can be converted to ATP

NTP Reactions

Activation of a reaction is generally a two step process - Pi or PPi or NMP (usually AMP ) is bound to substrate or enzyme - Phosphate containing moiety is displaced - Group may go to enzyme to substrate before being displaced In some instances ATP or GTP hydrolyzed directly-often to provide energy for movement ‒ ribosome movement on mRNA Phosphorylation of proteins can change conformation to cause activity ‒ Na+K+ ATPase transports ions using cycling of phosphorylation ATP or other high energy phosphate compounds can donate phosphates to NDPs to make NTPs 2 ADP can be used to make ATP + AMP ‒ When ATP low ‒ Can run in reverse when ATP high

Transport of ADP and Pi into the Matrix

Adenine nucleotide and phosphate translocases. Transport systems of the inner mitochondrial membrane carry ADP and Pi into the matrix and newly synthesized ATP into the cytosol. The adenine nucleotide translocase is an antiporter; the same protein moves ADP into the matrix and ATP out. The effect of replacing ATP4- with ADP3- in the matrix is the net efflux of one negative charge, which is favored by the charge difference across the inner membrane (outside positive). At pH 7, Pi is present as both HPO42- and H2PO4-; the phosphate translocase is specific for H2PO4- . There is no net flow of charge during symport of H2PO4- and H+, but the relatively low proton concentration in the matrix favors the inward movement of H+. Thus the proton-motive force is responsible both for providing the energy for ATP synthesis and for transporting substrates (ADP and Pi) into and product (ATP) out of the mitochondrial matrix. All three of these transport systems can be isolated as a single membranebound complex (ATP synthasome).

polyunsaturated fatty acids - first double bond requires isomerization ( starts at odd numbered C)

After isomerase converts cis to trans double bond, then the final 3 steps of β-oxidation can continue. The next round of β-oxidation starts with formation of the trans double bond at C2 but enoyl-CoA hydratase cannot work because of cis double bond at C4 Oxidation of a polyunsaturated fatty acid. The example here is linoleic acid, as linoleoyl-CoA (∆9,12). Oxidation requires a second auxiliary enzyme in addition to enoyl-CoA isomerase: NADPH-dependent 2,4-dienoyl-CoA reductase. The combined action of these two enzymes converts a trans-∆2,cis-∆4-dienoyl-CoA intermediate to the trans-∆2-enoyl-CoA substrate necessary for β oxidation.

several phosphorylated compounds have large negative deltaG' for hydrolysis

Again, electrostatic repulsion within the reactant molecule is relieved The products are stabilized via resonance or by more favorable solvation The product undergoes tautomerization Hydrolysis of phosphoenolpyruvate (PEP). Catalyzed by pyruvate kinase, this reaction is followed by spontaneous tautomerization of the product, pyruvate. Tautomerization is not possible in PEP, and thus the products of hydrolysis are stabilized relative to the reactants. Resonance stabilization of Pi also occurs

from pyruvate to phosphoenolpyruvate

Alternative paths from pyruvate to phosphoenolpyruvate The relative importance of the two pathways depends on the availability of lactate or pyruvate and the cytosolic requirements for NADH for gluconeogenesis The path on the right predominates when lactate is the precursor because cytosolic NADH is generated in the lactate dehydrogenase reaction and does not have to be shuttled out of the mitochondrion The requirements of ATP for Pyruvate carboxylase and GTP for PEP carboxykinase are omitted for simplicity *** NADH must be produced in cytosol to maintain NADH levels so gluconeogenesis continues to run -- needs NADH

b-oxidation in peroxisome

Animals use peroxisomes for breakdown of long chain or branched fatty acids Plants break down fatty acids mostly in peroxisomes and glyoxysomes Mitochondrial acyl-CoA dehydrogenase passes electrons from FADH2 into electron transport chain using Electron Transferring Flavoprotein - Energy captured as ATP Peroxisomal/glyoxysomal acyl-CoA oxidase passes electrons from FADH2 to molecular oxygen making hydrogen peroxide - Energy released as heat - Hydrogen peroxide eliminated by catalase

What types of fatty acids get broken down in peroxisomes in animals?

Animals use peroxisomes in order to breakdown long-chain or in some cases the branched fatty acids. However, some energy is lost when beta-oxidation is taken place within the peroxisome.

peripheral membrane proteins

Associate with the polar head groups on one side of membranes Relatively loosely associated with membrane - non-covalent interactions with the lipid head groups or aqueous domains of integral membrane proteins Can be removed by disrupting ionic/polar interactions either with high salt or change in pH Purified peripheral membrane proteins are no longer associated with any lipids

measurement of the standard reduction potential of a redox pair

At the standard hydrogen electrode (reference electrode), the half-reaction 2 H3O+ + 2 e- « 2 H2O + H2 takes place. At the other electrode, the half-reaction is as follows: Oxidized form + e- « reduced form. The combination of the oxidized and reduced version of a molecule is called a "conjugate redox pair". If the electron affinity of the oxidized form of the conjugate redox pair is higher than the electron affinity of H3O+, the standard reduction potential is positive. Otherwise, it is negative.

energy from reduced fuels is used to synthesize ATp in mitochondria of eukaryotes

Carbohydrates, lipids, and amino acids are the main reduced fuels for the cell Electrons from the oxidation of reduced fuels are transferred to produce reduced cofactors-NADH or FADH2 In oxidative phosphorylation, energy from NADH and FADH2 are used to make ATP In prokaryotes, this can be done using the plasma membrane if the prokaryote lives in an oxygenate environment

Oxidation of Propionyl-CoA

Carboxyl group addition needs energy > change stereoisomer D to L > Exchange groups to feed into citric acid cycle Oxidation of propionyl-CoA produced by β oxidation of odd-number fatty acids. The sequence involves the carboxylation of propionyl-CoA to D-methylmalonyl-CoA and conversion of the latter to succinyl-CoA. This conversion requires epimerization of D- to L-methylmalonyl-CoA, followed by a remarkable reaction in which substituents on adjacent carbon atoms exchange positions

energy flow in cellular respiration

Catabolism of proteins, fats, and carbohydrates in the three stages of cellular respiration. Stage 1: oxidation of fatty acids, glucose, and some amino acids yields acetyl-CoA. Stage 2: oxidation of acetyl groups in the citric acid cycle includes four steps in which electrons are abstracted. Stage 3: electrons carried by NADH and FADH2 are funneled into a chain of mitochondrial (or, in bacteria, plasma membrane-bound) electron carriers—the respiratory chain—ultimately reducing O2 to H2O. This electron flow drives the production of ATP. glucose -> NADH -> electron transport chain -> proton-motive force -> ATP

isocitrate dehydrogenase reaction

Catalyzes oxidative decarboxylation - Generate NADH - Lose a carbon as CO2-complete oxidation of carbon Oxidation converts an alcohol to a ketone - Transfers a hydride to NAD+ to make NADH Then carboxyl group on carbon 3 leaves as CO2 Highly thermodynamically favorable/irreversible -Regulated by product inhibition and ATP

Membrane composition is variable in different organelles Lipid composition of the plasma membrane and organelle membranes of fa rat hepatocyte The functional specialization of each membrane type is reflected in its unique lipid composition Cholesterol is prominent in plasma membranes but barely detectable in mitochondrial membranes Cardiolipin is a major component of the inner mitochondrial membrane but not of the plasma membrane Phosphatidylserine, phosphatidylinositol, and phosphatidylglycerol are relatively minor components of most membranes but serve critical functions Phosphatidylinositol and its derivatives, for example, are important in signal transduction triggered by hormones Spingolipids, phosphatidylcholine, and phosphatidylethanolamine are present in most membranes but in varying proportions Glycolipids, which are major components of the chloroplast membranes of plants, are virtually absent from animal cells

Cell membranes are asymmetric Every component of the membrane exhibits asymmetry Lipids Outer and inner leaflets have different lipid compositions Proteins Individual peripheral membrane proteins are only associated with one side of the membrane Integral membrane proteins have different domains on different sides of the membrane Specific lipid anchors added to proteins target the protein to a specific leaflet Carbohydrates Only on the outside of cells Membranes can be electrically polarized Plasma membrane-inside negative ~ -50mV Asymmetric distribution of phospholipids between the inner and outer monolayers of the erythrocyte plasma membrane The distribution of a specific phospholipid is determined by treating the intact cell with phospholipase C, which cannot reach lipids in the inner monolayer (leaflet) but removes the head groups of lipids in the outer monolayer The proportion of each head group released provides an estimate of the fraction of each lipid in the outer monolayer

transport across membranes

Cell membranes are permeable to small nonpolar molecules that passively diffuse through the membrane Passive diffusion of polar molecules involves desolvation and thus has a high activation energy barrier Transport across the membrane can be facilitated by proteins that provide an alternative diffusion path Such proteins are called transporters

structure of coenzyme A

Coenzymes are not a permanent part of the enzymes' structure. They associate, fulfill a function, and dissociate The function of CoA in this reaction is to accept and carry acetyl groups Coenzyme A (CoA). A hydroxyl group of pantothenic acid is joined to a modified ADP moiety by a phosphate ester bond, and its carboxyl group is attached to β-mercaptoethylamine in amide linkage. The hydroxyl group at the 3' position of the ADP moiety has a phosphoryl group not present in free ADP. The —SH group of the mercaptoethylamine moiety forms a thioester with acetate in acetyl-coenzyme A (acetyl-CoA) (lower left).

b-oxidation in mitochondria vs peroxisomes or glyoxysomes

Comparison of β oxidation in mitochondria and in peroxisomes and glyoxysomes. The peroxisomal/glyoxysomal system differs from the mitochondrial system in three respects: (1) the peroxisomal system prefers very-long-chain fatty acids; (2) in the first oxidative step electrons pass directly to O2, generating H2O2, and (3) the NADH formed in the second oxidative step cannot be reoxidized in the peroxisome or glyoxysome, so reducing equivalents are exported to the cytosol, eventually entering mitochondria. The acetyl-CoA produced by peroxisomes and glyoxysomes is also exported; the acetate from glyoxysomes (organelles found only in germinating seeds) serves as a biosynthetic precursor (see Fig. 17-15). Acetyl-CoA produced in mitochondria is further oxidized in the citric acid cycle.

summary of electron transport

Complex I -> Complex IV 1NADH + 11H+(N) + ½O2 ——> NAD+ + 10H+(P) + H2O deltaG' =-nFdeltaE' = -220kJ/mol •Complex II -> Complex IV FADH2 + 6H+(N) + ½O2 ——> FAD + 6H+(P) + H2O deltaG' =-nFdeltaE' = -150kJ/mol ( succinate to O2) Difference in number of protons transported means differences in number of ATP synthesized. 2.5 ATP per NADH and 1.5 ATP per FADH2

alpha-ketoglutarate a-ketoglutarate dehydrogenase reaction

Complex similar to pyruvate dehydrogenase - Same coenzymes, identical mechanisms - Active sites different to accommodate different-sized substrates A conserved mechanism for oxidative decarboxylation. The pathways shown employ the same five cofactors (thiamine pyrophosphate, coenzyme A, lipoate, FAD, and NAD+), closely similar multienzyme complexes, and the same enzymatic mechanism to carry out oxidative decarboxylations of pyruvate (by the pyruvate dehydrogenase complex), α-ketoglutarate (in the citric acid cycle), and the carbon skeletons of the three branched-chain amino acids, isoleucine (shown here), leucine, and valine. A fourth reaction, catalyzed by glycine decarboxylase, involves a very similar mechanism

citrate synthase reaction

Condensation of acetyl-CoA and oxaloacetate -only reaction with C-C bond formation Rate-limiting step of Citric Acid Cycle Activity largely depends on [oxaloacetate] Highly thermodynamically favorable/irreversible - Regulated by substrate availability and product inhibition

mitochondrial ATP synthase complex

Contains two functional units: F1 Soluble complex in the matrix Catalyzes the synthesis of ATP from ADP and Pi Fo Integral membrane complex Transports protons from IMS to matrix, dissipating the proton gradient Energy transferred to F1 to catalyze phosphorylation of ADP

transport of fatty acids requires conversion to fatty acyl-CoA

Conversion of a fatty acid to a fatty acyl-CoA. The conversion is catalyzed by fatty acyl-CoA synthetase and inorganic pyrophosphatase. Fatty acid activation by formation of the fatty acyl-CoA derivative occurs in two steps. The overall reaction is highly exergonic.

What is the mobile electron carrier that moves electrons between Complex III and IV? Where in the mitochondrion does it move?

Cytochrome c moves in the intermembrane space.

ketone bodies as fuel

D-β-Hydroxybutyrate as a fuel. D-β-Hydroxybutyrate, synthesized in the liver, passes into the blood and thus to other tissues, where it is converted in three steps to acetyl-CoA. It is first oxidized to acetoacetate, which is activated with coenzyme A donated from succinyl-CoA, then split by thiolase. The acetyl-CoA thus formed is used for energy production.

structure of a mitochondrion

Double membrane leads to four distinct compartments: 1. Outer Membrane: - Relatively porous membrane allows passage of metabolites 2. Intermembrane Space (IMS): - similar environment to cytosol - higher proton concentration (lower pH) 3. Inner Membrane - Relatively impermeable, with proton gradient across it - Location of electron transport chain complexes (including succinate dehydrogenase as part of Complex II) - Infolding of membrane produces cristae- serve to increase the surface area 4. Matrix - Location of the citric acid cycle and parts of lipid and amino acid metabolism - Lower proton concentration (higher pH)

Why does the cell need to have a process that reconverts NADH to NAD+ when no oxygen is available?

During anaerobic condition or extraneous work, the NAD+ must be regenerated in order for the glyceraldehyde-3P dehydrogenase to perform the oxidative phosphorylation and produce 1,3-bisphosphoglycerate, to keep glycolysis running. When there is no oxygen, cells must utilize fermentation for energy. The final fermentation reactions produce NAD+ to be used in glycolysis so the glycolysis pathway can continue to run. Without this, the supply of NAD+ would be depleted and glycolysis and ATP production would stop.

What happens to the fatty acid during each of the 2 redox reactions of β-oxidation? beta-oxidation

During the first redox reaction the transfer of 2 protons and 2 electrons from fatty acyl-CoA to FAD results in the formation of the trans double bond During the second redox reaction step 3, the hydroxyl group (alcohol) on beta carbon is oxidized to a ketone (hydroxyacyl to ketoacyl). Note: naturally occurring double bonds are cis but acyl-CoA dehydrogenase produces a trans double bond.

Why do E. coli change the composition of their membranes in response to temperature?

E. coli respond to temperature changes by changing their membrane composition because E. coli need to have a fluid membrane. So if the temperature decreases, more unsaturated fatty acids are needed in the membrane to keep it in the liquid-disordered state. On the other hand, if the temperature increases, more saturated fatty acids are needed.

electron transport chain complexes contain a series of electron carriers

Each complex contains multiple redox centers consisting of: - Flavin Mononucleotide ( FMN) or Flavin Adenine Dinucleotide ( FAD) Initial electron acceptors for Complex I and Complex II Can carry two electrons but can transfer one at a time - Cytochromes a, b or c - Iron-sulfur clusters and Heme groups - Copper centers and iron-copper centers

b-oxidation pathway

Each pass removes one acetyl moiety in the form of acetyl-CoA. The major pathway of fatty acid oxidation to produce NADH, FADH2, and acetyl coenzyme A. the process by which fatty acids are broken down to generate Acetyl-CoA which then enters the Citric Acid cycle. The β-oxidation pathway. (a) In each pass through this four-step sequence, one acetyl residue (shaded in pink) is removed in the form of acetyl-CoA from the carboxyl end of the fatty acyl chain—in this example palmitate (C16), which enters as palmitoyl-CoA. (b) Six more passes through the pathway yield seven more molecules of acetyl-CoA, the seventh arising from the last two carbon atoms of the 16-carbon chain. Eight molecules of acetyl-CoA are formed in all.

oxidative phosphorylation

Electrons from the reduced cofactors NADH and FADH2 are passed to proteins in the respiratory chain Oxygen is the ultimate electron acceptor for these electrons Energy of oxidation is used to phosphorylate ADP

NADH from glycolysis

Energy carrier for cellular respiration, makes 2 ATP at electron transport chain, because it loses energy traveling from the cytoplasm Must get from cytosol into mitochondria - Malate-aspartate shuttle electrons given to malate which has transporter into matrix where NADH is regenerated. Aspartate is transported out to cytosol NADH goes through Complex I-get 2.5 ATP for each NADH - Glycerol 3-P shuttle Electrons given to dihydroxyacetone phosphate to make glycerol-3P oxidation of glycerol-3P to dihydroxyacetone phosphate with electrons going to reduce FAD. Electrons transferred from FADH2 to ubiquinone-get 1.5 ATP for each FADH2

Polar solutes need alternative paths to cross cell membranes

Energy changes accompanying passage of a hydrophilic solute through the lipid bilayer of a biological membrane. (a) In simple diffusion, removal of the hydration shell is highly endergonic, and the energy of activation (∆G‡) for diffusion through the bilayer is very high. (b) A transporter protein reduces the ∆G‡ for transmembrane diffusion of the solute. It does this by forming noncovalent interactions with the dehydrated solute to replace the hydrogen bonding with water and by providing a hydrophilic transmembrane pathway.

proton transport linked to Chemical energy of ATP

Energy of ATP hydrolysis can be used to pump protons across the membrane against a gradient - pH control in the cell by F-type ATPase Energy of the proton gradient can be used to synthesize ATP - in chloroplast and mitochondrial membranes by ATP synthase

differences in peroxisomes and glyoxysomes

Enoyl-CoA hydratase and b-hydroxyacyl-CoA dehydrogenase activities occur in one enzyme-Multifunctional Protein In Animals, Acyl-CoA acetyltransferase (thiolase) is only active for long chain fatty acids. Once fatty acids are shortened, they are transported to mitochondria In glyoxysomes of plants, acetyl-CoA goes through the glyoxylate cycle Produces succinate - Transported to mitochondria to enter citric acid cycle - Precursor for gluconeogenesis - Link that allows conversion of fatty acids into hexoses

formation of ketone bodies

Entry of acetyl-CoA into citric acid cycle requires oxaloacetate When oxaloacetate is depleted, acetyl-CoA is converted into ketone bodies - Frees Coenzyme A for continued β-oxidation Ketone bodies are acetone, acetoacetate and β-hydroxybutyrate The first step is reverse of the last step in the b-oxidation- thiolase reaction joins two acetate units D-β-Hydroxybutyrate as a fuel. D-β-Hydroxybutyrate, synthesized in the liver, passes into the blood and thus to other tissues, where it is converted in three steps to acetyl-CoA. It is first oxidized to acetoacetate, which is activated with coenzyme A donated from succinyl-CoA, then split by thiolase. The acetyl-CoA thus formed is used for energy production.

sequence of events in oxidative decarboxylation of pyruvate

Enzyme 1 Step 1: Decarboxylation of pyruvate to an aldehyde Step 2: Oxidation of aldehyde to a thioester ‒ Lipoyl cofactor reduced and binds aldehyde to form the thioester Enzyme 2 Step 3: Formation of acetyl-CoA ( product 1) Enzyme 3 Step 4: Reoxidation of the lipoyl cofactor with reduction of FAD to FADH2 Step 5: Regeneration of the oxidized FAD cofactor forms reduced NADH ( product 2)

succinate dehydrogenase reaction

Enzyme integral to mitochondrial inner membrane - aka Complex II in the electron-transport chain Oxidation of the alkane to alkene requires reduction of FAD to FADH2 - Reduction potential of NAD is too low FAD is a prosthetic group - Because Succinate Dehydrogenase is integral to the mitochondrial inner membrane, FADH2 can participate in redox reactions in electron transport Near equilibrium/reversible - Product concentration kept low to pull forward

Why is everything done twice in the payoff phase?

Everything is done twice in the payoff phase because glucose is turned into two glyceraldehyde 3-phosphate molecules at the end of the preparatory phase, each of which go through the payoff phase.

redox reactions can be considered a combination of two half-reactions

Example for a complete redox reaction: ethanol + NAD+ « acetaldehyde + NADH + H+ (catalyzed by alcohol dehydrogenase) Half-reactions: (1) acetaldehyde + 2 H+ + 2 e- « ethanol (2) NAD+ + H+ + 2 e- « NADH Each of these reactions constitutes a conjugate redox pair with a standard reduction potential ( E' )

The Q Cycle

Experimentally, four protons are added to the intermembrane space per two electrons that reach Cytochrome c Two molecules of Ubiquinol (QH2) become oxidized - Two molecules of Cytochrome c become reduced- 1 electron from each QH2 goes to one Cytochrome c - Two protons come from each QH2- released into the intermembrane space One molecule of ubiquinone (Q) becomes reduced using 2 electrons from QH2 oxidation - Two protons from the matrix used to reduce Q Net reaction- four protons per (net one) ubiquinol (QH2) that binds to Complex III The Q cycle provides a good model that explains how two additional protons are picked up from the matrix The Q cycle, shown in two stages. The path of electrons through Complex III is shown by blue arrows. The movement of various forms of ubiquinone is shown with black arrows. In the first stage (left), Q on the N side is reduced to the semiquinone radical, which moves back into position to accept another electron. In the second stage (right), the semiquinone radical is converted to QH2. Meanwhile, on the P side of the membrane, two molecules of QH2 are oxidized to Q, releasing two protons per Q molecule (four protons in all) into the intermembrane space. Each QH2 donates one electron (via the Rieske Fe-S center) to cytochrome c1, and one electron (via cytochrome b) to a molecule of Q near the N side, reducing it in two steps to QH2. This reduction also consumes two protons per Q, which are taken up from the matrix (N side). Reduced cyt c1 passes electrons one at a time to cyt c, which dissociates and carries electrons to Complex IV.

Succinate Dehydrogenase aka Complex II

FAD accepts two electrons from succinate to make FADH2 Electrons are passed, one at a time, via iron-sulfur centers to ubiquinone, which becomes reduced (QH2) Does not transport protons across the membrane Structure of Complex II ( succinate dehydrogenase). (PDB ID 1ZOY) This complex (shown here is the porcine heart enzyme) has two transmembrane subunits, C and D; the cytoplasmic extensions contain subunits A and B. Just behind the FAD in subunit A is the binding site for succinate. Subunit B has three Fe-S centers, ubiquinone is bound to subunit B, and heme b is sandwiched between subunits C and D. Two phosphatidylethanolamine molecules are so tightly bound to subunit D that they show up in the crystal structure. Electrons move (blue arrows) from succinate to FAD, then through the three Fe-S centers to ubiquinone. The heme b is not on the main path of electron transfer but protects against the formation of reactive oxygen species (ROS) by electrons that go astray.

Where do FADH2 from the citric acid cycle and NADH enter the electron transport chain?

FADH2 enters at Complex II (succinate dehydrogenase) and NADH enters at Complex I.

breakdown of glycogen

FIRST enzyme - glycogen phosphorylate - breaks alpha1-4 a1-4 linkage between glycose molecules - uses inorganic phosphate instead of water to cleave the bond-phosphorolysis - cleaves until 4 glucoses remain before a branch point product of glucose 1-phosphate which cannot be transported out of the cell enzyme is regulated to only breakdown glycogen when ATP is needed SECOND enzyme - glycogen debranching enzyme - transfers 3 of the 4 remaining glucoses of one branch to another branch - cleaves off remaining glucose that is attached alpha1-6 a1-6 as glucose NOT glucose-1P THIRD enzyme - phosphoglucomutase - converts glucose 1-phosphate to glucose 6-phosphate which can feed in glycolysis in liver, glucose 6-phosphatase then cleaves off phosphate-glucose is transported out to the blood for other organs Debranching enzyme has two active sites each with critical Asp or Glu residues to break glucose attachments with their oxygens

fat storage in white adipose tissue

Fat stores in cells. (a) Cross section of human white adipose tissue. Each cell contains a fat droplet (white) so large that it squeezes the nucleus (stained red) against the plasma membrane.

fructose 1,6-bisP carbons in glyceraldehyde-3P

Fate of the glucose carbons in the formation of glyceraldehyde 3-phosphate (a) The origin of the carbons in the two threecarbon products of the aldolase and triose phosphate isomerase reactions The end product of the two reactions is glyceraldehyde 3-phosphate (two molecules) (b) Each carbon of glyceraldehyde 3-phosphate is derived from either of two specific carbons of glucose Note that the numbering of the carbon atoms of glyceraldehyde 3-phosphate differs from that of the glucose from which it is derived In glyceraldehyde 3-phosphate, the most complex functional group (the carbonyl) is specified as C-1 This numbering change is important for interpreting experiments with glucose in which a single carbon is labeled with a radioisotope

Fatty Acid Transport into Mitochondria

Fats are degraded into fatty acids and glycerol in the cytoplasm of adipocytes Fatty acids are transported to other tissues for fuel b-oxidation of fatty acids in animals mostly occurs in mitochondria Small (< 12 carbons) fatty acids diffuse freely across mitochondrial membranes Larger fatty acids (most free fatty acids) are transported via acyl-carnitine/carnitine transporter

acyl-carnitine / carnitine transporter

Fatty acid entry into mitochondria via the acyl-carnitine/carnitine transporter. After fatty acyl-carnitine is formed at the outer membrane or in the intermembrane space, it moves into the matrix by facilitated diffusion through the transporter in the inner membrane. In the matrix, the acyl group is transferred to mitochondrial coenzyme A, freeing carnitine to return to the intermembrane space through the same transporter. Acyltransferase I is inhibited by malonyl-CoA, the first intermediate in fatty acid synthesis (see Fig. 21-2). This inhibition prevents the simultaneous synthesis and degradation of fatty acids.

Fermentation includes glycolysis plus some additional reactions. What is the main role of these additional reactions?

Fermentation does not produce glucose. Maybe you are confusing it with gluconeogenesis The reactions of fermentation are necessary to convert NADH produced in glycolysis back to NAD+ so that glycolysis can continue. Without a pool of NAD+, glycolysis will stop at the Glyceraldenyde-3phosphate dehydrogenase step.

transverse movement of lipids

Flippases—cause the transverse movement of lipids Some flippases use energy of ATP to move lipids against the concentration gradient Motion of single phospholipids in a bilayer. (c) Three types of phospholipid translocaters in the plasma membrane. PE is phosphatidylethanolamine; PS is phosphatidylserine.

Fatty Acid Catabolism for Energy

For palmitate (16:0) - Repeating the above four-step process six more times (7 total) results in eight molecules of acetyl-CoA FADH2 is formed in each cycle ( 7 total) NADH is formed in each cycle (7 total) - Electrons from all FADH2 passed to Electron-transferring flavoprotein and eventually to ubiquinone in the electron transport chain - Electrons from NADH pass through Complex I to ubiquinone Acetyl-CoA enters citric acid cycle and is further oxidized into CO2 ( bypasses pyruvate dehydrogenase) - Makes GTP, 3 NADH, and FADH2

Synthesis of UDP-glucose-the substrate for glycogen synthase

Formation of a sugar nucleotide A condensation reaction occurs between a nucleoside triphosphate (NTP) and a sugar phosphate The negatively charged oxygen on the sugar phosphate serves as a nucleophile attacking the phosphate of the nucleoside triphosphate and displacing pyrophosphate The reaction is pulled in the forward direction by the hydrolysis of PPi ( 2Pi ) by inorganic pyrophosphatase

Cytochrome oxidase passes electrons to O2

Four Cytochome c that were reduced in Complex III bring electrons to Complex IV and are oxidized Four electrons are used to reduce one oxygen molecule to make two water molecules Four protons are picked up from the matrix in this process Four additional protons are pumped from the matrix to the intermembrane space

Write a reaction to form ATP using a GTP. (There is an → if you click on the © above the typing space. If you don't have 3 rows of buttons, you can expand the choices on the upper right corner)

GTP + ADP GDP + ATP

Glucose-6P is the starting material for this pathway. What other pathway is this used in?

Glucose-6P is also used in glycolysis also includes glycogen synthesis

glycerol from fats enters glycolysis

Glycerol kinase phosphorylates glycerol using ATP ‒ 2 Glycerol (3 C molecule) needed to be equivalent to glucose ‒ Would use 1 ATP/glycerol or 2/glucose equivalent Glycerol-3P dehydrogenase oxidizes glycerol-3P to dihydroxyacetone phosphate with reduction of NAD+ - DHAP feeds into glycolysis Subsequent reactions produce 2 ATP/glycerol (net 1 ATP) in glycolysis but more through citric acid cycle and oxidative phosphorylation Entry of glycerol into the glycolytic pathway.

Why is the action of glycogen phosphorylase called phosphorylysis?

Glycogen Phosphorylase is the enzyme that uses inorganic phosphate (not the H20) to cleave a glycosidic bond alpha 1-4 between glucose molecules that present in glycogen, therefore it called phosphorolysis.

dealing with branch points in glycogen

Glycogen breakdown near an (α1¦α6) branch point. Following sequential removal of terminal glucose residues by glycogen phosphorylase glucose residues near a branch are removed in a two-step process that requires a bifunctional debranching enzyme. First, the transferase activity of the enzyme shifts a block of three glucose residues from the branch to a nearby nonreducing end, to which they are reattached in (α1¦4) linkage The single glucose residue remaining at the branch point, in (α1¦6) linkage, is then released as free glucose by the debranching enzyme's (α1¦6) glucosidase activity The glucose residues are shown in shorthand form, which omits the —H, —OH, and —CH2OH groups from the pyranose rings *** leaves as glucose NOT glucose 1-P

What are the 2 actions of the glycogen debranching enzyme?

Glycogen debranching enzyme transfer 3 of the 4 glucoses to another branch and cleaves alpha 1-6 glycosidic bond.

glycogen is synthesized by glycogen synthase

Glycogen synthesis A glycogen chain is elongated by glycogen synthase The enzyme transfers the glucose residue of UDP glucose to the nonreducing end of a glycogen branch (see Fig. 7-13) to make a new ( a1-4 ) linkage.

Glycogenin starts a new glycogen chain

Glycogenin and the structure of the glycogen particle (a) Glycogenin catalyzes two distinct reactions Initial attack by the hydroxyl group of Tyr194 on C-1 of the glucosyl moiety of UDP-glucose results in a glucosylated Tyr residue The C-1 of another UDP-glucose molecule is now attacked by the C-4 hydroxyl group of the terminal glucose, and this sequence repeats to form a nascent glycogen molecule of eight glucose residues attached by (1¦4) glycosidic linkages.

What is the molecule that is the core of glycogen? What type of molecule is it?

Glycogenin is the protein at the core of glycogen. It is an enzyme that catalyzes the addition of the 1st 8 residues of glucose to itself using UDP-glucose.

Name the 3 enzymes that synthesize glycogen. Does each make α1,4 or α1,6 linkages between glucose monomers?

Glycogenin makes α1-4 connections, glycogen synthase makes α1-4 connections, and glycogen-branching enzyme makes α1-6 connections.

in eukaryotes citric acid cycle occurs in the mitochondria

Glycolysis occurs in the cytosol Citric acid cycle occurs in the mitochondrial matrix - Except succinate dehydrogenase, which is located in the inner membrane Oxidative phosphorylation occurs in the inner membrane

Glycolysis vs. Gluconeogenesis

Glycolysis occurs mainly in the muscle and brain Gluconeogenesis occurs mainly in the liver Glycolysis takes glucose and makes it into pyruvate Gluconeogenesis takes non-carb source (lactate, amino acid, or glycerol) and makes glucose Gluconeogenesis is basically reverse glycolysis except for 3 steps which are irreversible in glycolysis Gluconeogenesis used 4 alternates for this

The F1 catalyzes ADP + Pi <-> ATP

Hexamer arranged in three αβ dimers Dimers can exist in three different conformations: Open: empty - beta empty Loose: binding ADP and Pi - beta ADP Tight: catalyzes ATP formation and binds product - Beta ATP Mitochondrial ATP synthase complex. (b) (PDB ID 1BMF and PDB ID 1JNV) F1 viewed from above (that is, from the N side of the membrane), showing the three β (shades of purple) and three α (shades of gray) subunits and the central shaft (γ subunit, green). Each β subunit, near its interface with the neighboring α subunit, has a nucleotide-binding site critical to the catalytic activity. The single γ subunit associates primarily with one of the three αβ pairs, forcing each of the three β subunits into slightly different conformations, with different nucleotide-binding sites. In the crystalline enzyme, one subunit (β-ADP) has ADP (yellow) in its binding site, the next (β-ATP) has ATP (red), and the third (β-empty) has no bound nucleotide.

what does a hydropathy plot tell you?

Hydropathy plots suggest where transmembrane alpha helical segments occur in proteins. When a region of the protein is hydrophobic (positive hydropathy index) and roughly 20 amino acids in length, that region could be a transmembrane segment.

Why does the first bypass require 2 steps including the use of an ATP and a GTP?

If there was only 1 reaction, it might exist in cytosol. The reason why there has to be two reactions and the use of ATP and GTP is that the ΔG for the hydrolysis of the phosphate on phosphoenolpyruvate is a very high negative value. This means that to put the phosphate on pyruvate would be a highly unfavorable. In order to get enough free energy to do the reaction, there must be two steps, each using an NTP. Just for your knowledge-The first reaction puts a carboxyl group on the pyruvate and then the second reaction removes the carboxyl group (negative ΔG) and removes a phosphate from GTP (negative ΔG) to provide enough free energy to put the phosphate on to produce phosphoenolpyruvate (positive ΔG).

Glucose -6 phosphate can go to glycolysis or glycogen synthesis. What is the product of the reaction that utilizes glucose-6P for each pathway and what enzyme catalyzes the reaction? a) for glycolysis b) for glycogen synthesis

In glycolysis glucose-6P converted to fructose 6P by phosphohexose isomerase In glycogen synthesis glucose 6P is converted to glucose 1P by the enzyme phosphoglucomutase

What is the difference between passive and active transport?

In passive transport, the molecules/ions are being moved in the favorable direction (down their concentration and/or charge gradient), hence there is no need for added energy. In active transport, the molecules/ions are being moved in an unfavorable direction (up their concentration and/or charge gradient) so there needs to be an input of energy in order to make the transport occur. Remember that for charged molecules, both the concentration and the charge gradient play a role. For uncharged molecules, only the concentration difference matters.

6 types of integral membrane proteins formed by a helices alpha helices

Integral membrane proteins. For known proteins of the plasma membrane, the spatial relationships of protein domains to the lipid bilayer fall into six categories. Types I and II have a single transmembrane helix; the amino-terminal domain is outside the cell in type I proteins and inside in type II. Type III proteins have multiple transmembrane helices in a single polypeptide. In type IV proteins, transmembrane domains of several different polypeptides assemble to form a channel through the membrane. Type V proteins are held to the bilayer primarily by covalently linked lipids (see Fig. 11-15), and type VI proteins have both transmembrane helices and lipid anchors. In this figure, and in figures throughout the book, we represent transmembrane protein segments in their most likely conformations: as α helices of six to seven turns. Sometimes these helices are shown simply as cylinders. As relatively few membrane protein structures have been deduced by x-ray crystallography, our representation of the extramembrane domains is arbitrary and not necessarily to scale.

integral membrane proteins involved in movement of molecules across membranes

Ion Channels - Passive transport - Molecules move down their concentration gradient at rates close to diffusion - Generally don't become saturated Transporters - Can perform active or passive transport - Move molecules slower than diffusion - Can move molecules up their concentration gradient - Can be saturated

oxidation of double bonds starting at odd numbered carbons

Isomerase replaces action of acyl-CoA dehydrogenase for that cycle, resulting in 1 fewer FADH2. Oxidation of a monounsaturated fatty acid. Oleic acid, as oleoyl-CoA (∆9), is the example used here. Oxidation requires an additional enzyme, enoyl-CoA isomerase, to reposition the double bond, converting the cis isomer to a trans isomer, a normal intermediate in β oxidation.

aconitase reaction

Isomerizes molecule by removing and then adding H2O - Elimination of H2O from citrate gives a cis C=C bond - Addition of H2O to cis-aconitate is stereospecific - Citrate, a tertiary alcohol, is a poor substrate for oxidation - Isocitrate, a secondary alcohol, is a good substrate for oxidation because there are two hydrogens (one on the alcohol and one on the carbon the alcohol is attached to) to remove Thermodynamically unfavorable/reversible - Product concentration kept low to pull forward

What does UDP-glucose-4-epimerase produce?

It flips the OH on C-4 to make UDP-glucose

Why is it important to make NADH in cytosol during this bypass?

It is important for NADH to be made in the cytosol because levels need to be maintained in order for gluconeogenesis to keep running. Remember that glyceraldehyde-3P dehydrogenase is running in reverse in gluconeogenesis so it needs NADH as a substrate.

Why does the non-oxidative phase need to run twice to be complete?

It's correct to start with the non-oxidative phase is there to produce five glucose-6P. However, you need to add that in each round, it makes two 6 carbon sugars and one 3 carbon sugar. By running it two times, two 3 carbon sugars are made which can be combined to make another glucose-6P.

What are the products of the extra reactions for lactic acid fermentation? How many reactions are involved?

Lactate and NAD+ are the products of the lactic acid fermentation, it's one redox reaction that reduces pyruvate to lactate, by oxidizing NADH to NAD+. Lactate can be converted to glucose in the liver.

What is the enzyme that breaks down triacylglycerol into glycerol and 3 fatty acids?

Lipase breaks down triacylglercols into three fatty acids and glycerol.

hydropathy plots can predict helical transmembrane domains

Lipid composition of the plasma membrane and organelle membranes of a rat hepatocyte. The functional specialization of each membrane type is reflected in its unique lipid composition. Cholesterol is prominent in plasma membranes but barely detectable in mitochondrial membranes. Cardiolipin is a major component of the inner mitochondrial membrane but not of the plasma membrane. Phosphatidylserine, phosphatidylinositol, and phosphatidylglycerol are relatively minor components of most membranes but serve critical functions; phosphatidylinositol and its derivatives, for example, are important in signal transductions triggered by hormones. Sphingolipids, phosphatidylcholine, and phosphatidylethanolamine are present in most membranes but in varying proportions. Glycolipids, which are major components of the chloroplast membranes of plants, are virtually absent from animal cells.

Why do lipids produce more ATP when they are broken down?

Lipids produce more ATP when broken down because their carbons are more reduced compared to those of polysaccharides. Lipids usually have more carbons as well so more carbons to oxidize.

malate dehydrogenase reaction

Malate + NAD⁺ <--> Oxaloacetate + NADH + H⁺ Final step of the cycle Alcohol on Malate oxidized to a ketone and NAD+ is reduced to NADH Regenerates oxaloacetate for citrate synthase Highly thermodynamically UNfavorable/reversible - Oxaloacetate concentration kept VERY low by citrate synthase Pulls the reaction forward

cytochrome oxidase aka Complex IV

Mammalian cytochrome oxidase is a membrane protein with 13 subunits Contains two heme groups: a and a3 Contains copper ions - CuA: two ions that accept electrons from Cyt c - CuB: bonded to heme a3 forming a binuclear center that transfers four electrons to oxygen Structure of cytochrome oxidase (Complex IV). This complex from bovine mitochondria has 13 subunits, but only four core proteins are shown here (PDB ID 1OCC). (a) Complex IV, with four subunits in each of two identical units of a dimer. Subunit I (yellow) has two heme groups, a and a3, near a single copper ion, CuB (not visible here). Heme a3 and CuB form a binuclear Fe-Cu center. Subunit II (purple) contains two Cu ions complexed with the —SH groups of two Cys residues in a binuclear center, CuA, that resembles the 2Fe-2S centers of iron-sulfur proteins. This binuclear center and the cytochrome c-binding site are located in a domain of subunit II that protrudes from the P side of the inner membrane (into the intermembrane space). Subunit III (blue) is essential for rapid proton movement through subunit II. The role of subunit IV (green) is not yet known. (b) The binuclear center of CuA. The Cu ions (green spheres) share electrons equally. When the center is reduced, the ions have the formal charges Cu1+Cu1+; when oxidized, Cu1.5+Cu1.5+. Six amino acid residues are ligands around the Cu ions: two His, two Cys, Glu, and Met.

oxidation reduction reactions

Many biochemical oxidation-reduction reactions involve transfer of two electrons In order to keep charges in balance, proton transfer often accompanies electron transfer In many dehydrogenases, the reaction proceeds by a stepwise transfers of proton (H+) and hydride (:H-) An oxidation-reduction reaction. Shown here is the oxidation of lactate to pyruvate. In this dehydrogenation, two electrons and two hydrogen ions (the equivalent of two hydrogen atoms) are removed from C-2 of lactate, an alcohol, to form pyruvate, a ketone. In cells the reaction is catalyzed by lactate dehydrogenase and the electrons are transferred to the cofactor nicotinamide adenine dinucleotide (NAD). This reaction is fully reversible; pyruvate can be reduced by electrons transferred from the cofactor.

membrane fusion

Many cellular functions especially in eukaryotes require membranes to fuse Membranes can fuse with each other without losing continuity Fusion can be spontaneous or protein-mediated Proteins can - bend membranes to form a vesicle - bring a vesicle close enough to fuse with a membrane Examples of protein-mediated fusion - Entry of influenza virus into the host cell - Release of neurotransmitters at nerve synapses

the cori cycle

Metabolic cooperation between skeletal muscle and the liver: the Cori cycle. Extremely active muscles use glycogen as energy source, generating lactate via glycolysis During recovery, some of this lactate is transported to the liver and converted to glucose via gluconeogenesis This glucose is released to the blood and returned to the muscles to replenish their glycogen stores The overall pathway (glucose → lactate → glucose) constitutes the Cori cycle.

oxidation of fatty acids with an odd number of carbons

Most dietary fatty acids have an even number of carbons Many plants and some marine organisms also synthesize fatty acids with an odd number of carbons Propionyl-CoA (last 3 carbons) forms from b-oxidation of fatty acids with an odd number of carbons Bacterial metabolism in the rumen of ruminants also produces propionyl-CoA

For the following reaction malate + NAD+ → oxaloacetate + NADH + H+ 1. Name the two conjugate redox pairs as Oxidized species/Reduced species 2. Write half reactions for each pair. (the → symbol can be found when you click on © in the text choices above the area where you will type your answer) 3. Which conjugate redox pair is getting oxidized in the original equation? 4. What is the deltaE' ΔEº' for this reaction? please show your work 5. What is the deltaG' ΔG º' for this reaction? please show your work

NAD+ is in (ox-state, due to no Hydrogen), NADH gaining hydrogen, thus reduced (red-state). Oxaloacetate + (2H+) + 2electrons= malate = -0.166V Since the reaction is started from malate (red-state), which is in reverse direction, thus the E will be +0.166V NAD+/NADH (electron acceptor)= -0.32V malate/oxaloacetate (electron donor) ΔE°'= −0.32 − (−0.166)= −0.154V ΔG º'= -nF(-0.154) = -2*96.5*(-0.154) = 29.722kJ/mole (not favorable) 2. NAD+ + H+ + 2e- → NADH and oxaloacetate + 2H+ + 2e- → malate 1. NAD+/NADH reduced species, Malate/oxaloacetate oxidized species 3. malate/oxaloacetate

Complex I

NADH : ubiquinone oxidoreductase (Complex I). NADH-ubiquinone, NADH:ubiquinone (PDB ID 3M9S) Complex I (the crystal structure from the bacterium Thermus thermophilus is shown) catalyzes the transfer of a hydride ion from NADH to FMN, from which two electrons pass through a series of Fe-S centers to the Fe-S center N-2 in the matrix arm of the complex. Electron transfer from N-2 to ubiquinone on the membrane arm forms QH2, which diffuses into the lipid bilayer. This electron transfer also drives the expulsion from the matrix of four protons per pair of electrons. The detailed mechanism that couples electron and proton transfer in Complex I is not yet known, but probably involves a Q cycle similar to that in Complex III in which QH2 participates twice per electron pair (see Fig. 19-12). Proton flux produces an electrochemical potential across the inner mitochondrial membrane ( N side negative, P side positive).

Why does it make sense that high [NADH] and [ATP] would allosterically downregulate certain Citric Acid Cycle enzymes?

NADH and ATP are the main end products of the citric acid cycle. If there is an excess of these end products, it does not make sense for the cell to continue to run this process and make these products. Therefore, it makes sense that high concentrations of NADH and ATP would downregulate the citric acid cycle, as it would prevent the cell from putting in unneeded work.

Oxidation of unsaturated fatty acids

Naturally occurring unsaturated fatty acids contain cis double bonds-very common in eukaryotes - NOT a substrate for enoyl-CoA hydratase of Step 2 Two additional enzymes are required - Isomerase: converts cis or trans double bonds starting at carbon 3 to trans double bonds at carbon 2 - Reductase: reduces cis double bonds not at carbon 3 Fatty acids with double bonds starting at odd number carbons require the isomerase Fatty acids with double bonds starting at even number fatty acids require both enzymes

conversion of pyruvate to Acetyl-CoA

Needed for pyruvate from glycolysis to enter Citric Acid Cycle- not needed for fatty acid catabolism Result of Reaction: - Oxidative decarboxylation of pyruvate - First carbons of glucose to be fully oxidized Catalyzed by the pyruvate dehydrogenase complex - Requires 5 cofactors - TPP, lipoyllysine, and FAD are prosthetic groups - NAD+ and CoA-SH are coenzymes Overall reaction catalyzed by the pyruvate dehydrogenase complex. The five coenzymes participating in this reaction, and the three enzymes that make up the enzyme complex, are discussed in the text.

In summary glycolysis:

Net Reaction: glucose + 2 NAD+ + 2 ADP + 2 Pi > 2 pyruvate + 2 NADH + 2 H+ + 2 ATP Used: 1 glucose 2 ATP 2 NAD+ 4 ADP 2 Pi Made: 2 pyruvate 4 ATP ( net of 2 ATP ) - used for energy requiring processes within the cell 2 NADH - must be reoxidized to NAD+ in order for glycolysis to continue , can be used to make ATP through oxidative phosphorylation glycolysis is heavily regulated - ensure proper use of nutrients - ensure production of ATP only when needed deltaG' = -146 kJ/mol for breakdown of glucose and +61 kJ/mol for the synthesis of 2 ATP

NAD+ and NADP+ are common redox cofactors

Nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate Coenzymes-can dissociate from the enzyme after the reaction and react elsewhere to return to original redox state In a typical biological oxidation reaction, hydride from an alcohol is transferred to NAD+ giving NADH NADPH usually used to reduce other molecules NAD+ and NADP+ work in different reactions NAD and NADP. (a) Nicotinamide adenine dinucleotide, NAD+, and its phosphorylated analog NADP+ undergo reduction to NADH and NADPH, accepting a hydride ion (two electrons and one proton) from an oxidizable substrate The hydride ion is added to either the front (the A side) or the back (the B side) of the planar nicotinamide ring

non-oxidative phase regenerates Glucose-6-P from Ribose-5-P used in tissues requiring more NADPH than Ribose-5-P ( liver and adipose tissue )

Nonoxidative reactions of the pentose phosphate pathway. (a) These reactions convert pentose phosphates to hexose phosphates, allowing the oxidative reactions (see Fig. 14-22) to continue. Transketolase and transaldolase are specific to this pathway; the other enzymes also serve in the glycolytic or gluconeogenic pathways. (b) A schematic diagram showing the pathway from six pentoses (5C) to five hexoses (6C). Note that this involves two sets of the interconversions shown in (a). Every reaction shown here is reversible; unidirectional arrows are used only to make clear the direction of the reactions during continuous oxidation of glucose 6-phosphate. In the light-independent reactions of photosynthesis, the direction of these reactions is reversed (see Fig. 20-10).

In the Q cycle in Complex III, where do the 2 electrons from the oxidation of each ubiquinol ( QH2 ) end up?

One electron from the oxidation of ubiquinol goes to reduce cytochrome c , and the other goes to reduce ubiquinone.

NADH-ubiquinone oxidoreductase aka Complex I

One of the largest macro-molecular assemblies in the mammalian cell Over 40 different polypeptide chains, encoded by both nuclear and mitochondrial genes NADH binding site on the matrix side of the membrane Noncovalently bound flavin mononucleotide (FMN) accepts two electrons from NADH but passes them on one at a time Several iron-sulfur centers pass one electron at a time toward the ubiquinone binding site Final reduction is ubiquinone to ubiquinol (Q to QH2)

some membrane proteins made of beta sheets b-sheets b sheets

Only found in bacteria, mitochondria and chloroplasts H-bonding maximized by circle of β strands Membrane proteins with -barrel structure. Three proteins of the E. coli outer membrane are shown, viewed in the plane of the membrane. FepA (PDB ID 1FEP), involved in iron uptake, has 22 membranespanning β strands. OmpLA (derived from PDB ID 1QD5), a phospholipase, is a 12-stranded β barrel that exists as a dimer in the membrane. Maltoporin (derived from PDB ID 1MAL), a maltose transporter, is a trimer; each monomer consists of 16 β strands.

second starts at even numbered C - requires reduction / isomerization

Oxidation of a polyunsaturated fatty acid. The example here is linoleic acid, as linoleoyl-CoA (∆9,12). Oxidation requires a second auxiliary enzyme in addition to enoyl-CoA isomerase: NADPH-dependent 2,4-dienoyl-CoA reductase. The combined action of these two enzymes converts a trans-∆2,cis-∆4-dienoyl-CoA intermediate to the trans-∆2-enoyl-CoA substrate necessary for β oxidation.

overall reaction of pyruvate dehydrogenase

Oxidative decarboxylation of pyruvate to acetyl-CoA by the PDH complex. The fate of pyruvate is traced in red. In step 1 pyruvate reacts with the bound thiamine pyrophosphate (TPP) of pyruvate dehydrogenase (E1), undergoing decarboxylation to the hydroxyethyl derivative Pyruvate dehydrogenase also carries out step 2, the transfer of two electrons and the acetyl group from TPP to the oxidized form of the lipoyllysyl group of the core enzyme, dihydrolipoyl transacetylase (E2), to form the acetyl thioester of the reduced lipoyl group Step 3 is a transesterification in which the —SH group of CoA replaces the —SH group of E2 to yield acetyl-CoA and the fully reduced (dithiol) form of the lipoyl group. step 4 dihydrolipoyl dehydrogenase (E3) promotes transfer of two hydrogen atoms from the reduced lipoyl groups of E2 to the FAD prosthetic group of E3, restoring the oxidized form of the lipoyllysyl group of E2 step 5 the reduced FADH2 of E3 transfers a hydride ion to NAD+, forming NADH The enzyme complex is now ready for another catalytic cycle

Why must oxidative phosphorylation take place within an impermeable membrane?

Oxidative phosphorylation must take place within an impermeable membrane because an electrochemical proton gradient needs to be formed. If protons could diffuse across the membrane, this gradient would not be able to be established.

Why can phosphoenolpyruvate (PEP) give a phosphate group to ADP to make ATP?

PEP has a more negative free energy for the hydrolysis of the phosphate than does ATP. I know it's a pain to write it all out, but the reaction we are talking about matters. Phosphoenolpyruvate PEP can donate a phosphate to ADP because PEP has a more negative standard free energy compared to ATP. Therefore, PEP can donate its phosphate group to ADP and it is a favorable reaction.

Transport across membranes

Passive transport across a membrane must be energetically favorable. - Concentration dependence: The solute moves toward equilibrium across the membrane. - Electrochemical dependence: The solute moves toward charge equilibrium across the membrane. If not energetically favorable, energy must be added to the system to achieve transport- active transport Movement of solutes across a permeable membrane. (a) Net movement of an electrically neutral solute is toward the side of lower solute concentration until equilibrium is achieved. The solute concentrations on the left and right sides of the membrane, as shown here, are designated C1 and C2. The rate of transmembrane solute movement (indicated by the arrows) is proportional to the concentration ratio. (b) Net movement of an electrically charged solute is dictated by a combination of the electrical potential (Vm) and the ratio of chemical concentrations (C2/C1) across the membrane; net ion movement continues until this electrochemical potential reaches zero.

model for glucose transport

Passive transport of glucose into the cell. If [glucose]in = [glucose]out, transport stops. Glucose metabolism tends to make this unlikely. Model of glucose transport into erythrocytes by GLUT1. The transporter exists in two conformations: T1, with the glucose-binding site exposed on the outer surface of the plasma membrane, and T2, with the binding site exposed on the inner surface. Glucose transport occurs in four steps. 1 Glucose in blood plasma binds to a stereospecific site on T1; this lowers the activation energy for 2 a conformational change from glucoseout • T1 to glucosein • T2, effecting the transmembrane passage of the glucose. 3 Glucose is released from T2 into the cytoplasm, and 4 the transporter returns to the T1 conformation, ready to transport another glucose molecule.

Flow of Electrons from Biological Fuels into the Electron Transport Chain

Path of electrons from NADH, succinate, fatty acyl-CoA, and glycerol 3-phosphate to ubiquinone. Ubiquinone (Q) is the point of entry for electrons derived from reactions in the cytosol, from fatty acid oxidation, and from succinate oxidation (in the citric acid cycle). Electrons from NADH pass through a flavoprotein with the cofactor FMN to a series of Fe-S centers (in Complex I) and then to Q. Electrons from succinate pass through a flavoprotein with the cofactor FAD and several Fe-S centers (in Complex II) on the way to Q. Glycerol 3-phosphate donates electrons to a flavoprotein (glycerol 3-phosphate dehydrogenase) on the outer face of the inner mitochondrial membrane, from which they pass to Q. Acyl-CoA dehydrogenase (the first enzyme of β oxidation) transfers electrons to electron-transferring flavoprotein (ETF), from which they pass to Q via ETF : ubiquinone oxidoreductase.

flow of electrons from biological fuels into the electron transport chain

Path of electrons from NADH, succinate, fatty acyl-CoA, and glycerol 3-phosphate to ubiquinone. Ubiquinone (Q) is the point of entry for electrons derived from reactions in the cytosol, from fatty acid oxidation, and from succinate oxidation (in the citric acid cycle). Electrons from NADH pass through a flavoprotein with the cofactor FMN to a series of Fe-S centers (in Complex I) and then to Q. Electrons from succinate pass through a flavoprotein with the cofactor FAD and several Fe-S centers (in Complex II) on the way to Q. Glycerol 3-phosphate donates electrons to a flavoprotein (glycerol 3-phosphate dehydrogenase) on the outer face of the inner mitochondrial membrane, from which they pass to Q. Acyl-CoA dehydrogenase (the first enzyme of β oxidation) transfers electrons to electron-transferring flavoprotein (ETF), from which they pass to Q via ETF : ubiquinone oxidoreductase. Step 3: dehydrogenation of alcohol Catalyzed by b-hydroxyacyl-CoA dehydrogenase Alcohol is oxidized to a ketone The enzyme reduces NAD+ to NADH Short fatty acyl chains catalyzed by soluble enzyme Long fatty acyl chains catalyzed by Trifunctional protein in inner membrane Step 4: release of acetyl-CoA Catalyzed by acyl-CoA acetyltransferase (thiolase) via covalent mechanism - The carbonyl carbon in b-ketoacyl-CoA is electrophilic - Active site thiol group (from cys) on the protein acts as nucleophile and releases acetyl-CoA - Terminal sulfur in Coenzyme A acts as nucleophile and picks up the fatty acid chain from the enzyme The net reaction is thiolysis of carbon-carbon bond (our third version of lysis) Short fatty acyl chains catalyzed by soluble enzyme Long fatty acyl chains catalyzed by Trifunctional protein

electron flow through Complex IV

Path of electrons through Complex IV. The three proteins critical to electron flow are subunits I, II, and III. The larger green structure includes the other 10 proteins in the complex. Electron transfer through Complex IV begins with cytochrome c (top). Two molecules of reduced cytochrome c each donate an electron to the binuclear center CuA. From here electrons pass through heme a to the Fe-Cu center (heme a3 and CuB). Oxygen now binds to heme a3 and is reduced to its peroxy derivative (O22-; not shown here) by two electrons from the Fe-Cu center. Delivery of two more electrons from cytochrome c (top, making four electrons in all) converts the O22- to two molecules of water, with consumption of four "substrate" protons from the matrix. At the same time, four protons are pumped from the matrix by an as yet unknown mechanism.

differences in reduction potential between two redox pairs ( deltaE' ) translate into a difference in free energy ( deltaG' )

Per definition, ΔE°' values are calculated as E°' of the electron acceptor minus E°' of the electron donor. Can also think of it as E of pair that goes from ox to red - E of pair that goes from red to ox Example: Pyruvate + NADH + H+ ® lactate + NAD+ E'° for the pyruvate/lactate pair: -0.185 V E'° for the NAD+/NADH pair: -0.32 V ∆E°' = E°'(e- acceptor) - E°'(e- donor) ΔE'° = -0.185 V - (-0.32 V) = 0.135 V Formula to calculate ΔG°': ΔG°' = -nF ΔE°' ΔG°' = -2 ´ 96.5 kJ/V·mol´ 0.135 V = -26.1 kJ/mol Whereas the standard reduction potential is a constant, the actual reduction potential of a redox pair is dependent on the relative concentrations of the reduced and oxidized species. This concentration dependence for a conjugate redox pair is as follows: E = E°' + RT/nF ln[electron acceptor]/[electron donor] E = E°' + RT/nF ln[Aox]/[Ared] R is the gas constant (0.00832 kJ/mol·K) F is the Faraday constant (96.5 kJ/V·mol) T is the absolute temperature (in Kelvin) n is the number of electrons transferred Example: Mixture of 0.1 M lactate and 1 mM pyruvate (conjugate redox pair) at pH 7 and 298°K: pyruvate + 2 H+ + 2 e- « lactate E = E'° + (lactate is the reduced species-has gained electrons compared to pyruvate) E = (- 0.185 V) + = - 0.245 V The protons are disregarded because the pH corresponds to biochemical standard conditions.

differences in standard free energy of hydrolysis of a phosphate

Phosphate can be transferred from compounds with more negative ΔG' to those with less negative ΔG' Reactions such as PEP + ADP → Pyruvate + ATP are favorable, and can be used to synthesize ATP Ranking of biological phosphate compounds by standard free energies of hydrolysis. This shows the flow of phosphoryl groups, represented by P, from high-energy phosphoryl group donors via ATP to acceptor molecules (such as glucose and glycerol) to form their low-energy phosphate derivatives. (The location of each compound's donor phosphoryl group along the scale approximately indicates the deltaG' of hydrolysis.) This flow of phosphoryl groups, catalyzed by kinases, proceeds with an overall loss of free energy under intracellular conditions. Hydrolysis of low-energy phosphate compounds releases Pi, which has an even lower phosphoryl group transfer potential (as defined in the text).

What enzymes catalyze the reactions that utilize the molecules in 3a as substrates?

Phosphoglycerate kinase uses 1,3 biphosphoglycerate as a substrate to form 3-phosphoglycerate Pyruvate kinase uses phosphoenolpyruvate as a substrate to form pyruvate.

The first step in glycolysis is phosphorylation of glucose. Besides metabolic pathways, why is this important to the functioning of the cell?

Phosphorylation of glucose is important for the cell because when a phosphate is added to a glucose, it prevents it from going back across the membrane through its transporter. So glucose is trapped now in the cell for energy.

ATP often provides energy to reactions in a two step process. In the first step, what can be bound to the enzyme or the substrate? (Hint, there are 3 answers. Feel free to give 1 at a time)

Pi, PPi or NMP (mostly AMP).

Regulation of Oxidative Phosphorylation

Primarily regulated by substrate availability - NADH or FADH2 and ADP/Pi - Due to coupling of the two reactions, substrates required for both electron transport and ATP synthesis Inhibition of OxPhos leads to accumulation of NADH - Inhibitor for enzymes in glycolysis and citric acid cycle Inhibitor of F1 ( IF1) - ATP synthase can work in reverse when there is no proton gradient - IF1 works in matrix-prevents hydrolysis of ATP during low oxygen by preventing ATPase from turning backwards - Only active at lower pH, encountered when electron transport it stalled (low oxygen) The rates of the citric acid cycle and oxidative phosphorylation are very close, because the products of the citric acid cycle feed into the ETC, and therefore oxidative phosphorylation

cellular respiration

Process in which cells consume O2 and produce CO2 Provides more energy (ATP) from glucose than glycolysis Also captures energy stored in lipids and amino acids Used by animals, plants, and many microorganisms Occurs in three major stages: - acetyl CoA production - acetyl CoA oxidation - electron transfer and oxidative phosphorylation

trisfunctional protein

Processes fatty acid chains with 12 or more carbons Performs last 3 steps of β oxidation Hetero-octamer - Four a subunits enoyl-CoA hydratase activity b-hydroxyacyl-CoA dehydrogenase activity - Four b subunits long-chain acyl-Co acetyltransferase (thiolase) activity May allow substrate channeling Associated with inner mitochondrial membrane Shorter chains including products of Trifunctional protein of less than 12 carbons processed by soluble enzymes in the matrix

liver is the source of ketone bodies

Production of ketone bodies increases during starvation (and diabetes) Ketone bodies are released by liver to bloodstream Acetoacetate and β-hydroxybutryrate can be broken down into 2 acetyl-CoA in other tissues including brain and muscle. Acetone can be converted to pyruvate. Organs other than liver can use ketone bodies as fuels

one turn of the citric acid cycle

Products of one turn of the citric acid cycle. At each turn of the cycle, three NADH, one FADH2, one GTP (or ATP), and two CO2 are released in oxidative decarboxylation reactions. most of the reactions are reversible

What kind of fatty acids gets broken down to proprionyl-CoA? What does proprionyl-CoA get converted to, and what pathway does this product go to?

Propionyl-CoA (has 3 carbons) forms by beta-oxidation of fatty acids with an odd number of carbons instead of usual ones which have even number of carbons. Propionyl-CoA can be oxidized to the Succinyl-CoA and can be fed into the citric acid cycle.

binding-change model

Proposed mechanism for the action of ATP synthase, involving concerted conformational changes, rotational motions, catalysis, and substrate binding and release. Binding-change model for ATP synthase. The F1 complex has three nonequivalent adenine nucleotide-binding sites, one for each pair of α and β subunits. At any given moment, one of these sites is in the β-ATP conformation (which binds ATP tightly), a second is in the β-ADP (loose-binding) conformation, and a third is in the β-empty (very-loose-binding) conformation. The proton-motive force causes rotation of the central shaft—the γ subunit, shown as a green arrowhead—which comes into contact with each αβ subunit pair in succession. This produces a cooperative conformational change in which the β-ATP site is converted to the β-empty conformation, and ATP dissociates; the β-ADP site is converted to the β-ATP conformation, which promotes condensation of bound ADP + Pi to form ATP; and the β-empty site becomes a β-ADP site, which loosely binds ADP + Pi entering from the solvent. This model, based on experimental findings, requires that at least two of the three catalytic sites alternate in activity; ATP cannot be released from one site unless and until ADP and Pi are bound at the other.

structure of integral membrane proteins

Proteins made of helices need helices of about 20 amino acids to cross the membrane - Amino acids must be hydrophobic if interacting with membrane - Amino acids interacting with other helices or central to a pore can be hydrophilic Proteins made of β sheet only need 7-9 amino acids in a strand to cross membrane - Amino acids with R groups pointing towards membrane must be hydrophobic - Amino acids with R group pointing into barrel may be hydrophilic Connections or domains extending out of the membrane can be hydrophilic

cytochromes

Proteins or subunits of complexes containing heme prosthetic groups One electron carriers Iron coordinating porphoryin ring derivatives a, b or c differ by ring additions Can be mobile or stationary Prosthetic groups of cytochromes. (a) Each group consists of four five-membered, nitrogen-containing rings in a cyclic structure called a porphyrin. The four nitrogen atoms are coordinated with a central Fe ion, either Fe2+ or Fe3+. Iron protoporphyrin IX is found in b-type cytochromes and in hemoglobin and myoglobin (see Fig. 4-17). Heme c is covalently bound to the protein of cytochrome c through thioether bonds to two Cys residues. Heme a, found in a-type cytochromes, has a long isoprenoid tail attached to one of the five-membered rings. The conjugated double-bond system (shaded light red) of the porphyrin ring has delocalized π electrons that are relatively easily excited by photons with the wavelengths of visible light, which accounts for the strong absorption by hemes (and related compounds) in the visible region of the spectrum. (b) Absorption spectra of cytochrome c (cyt c) in its oxidized (blue) and reduced (red) forms. The characteristic α, β, and γ bands of the reduced form are labeled.

Coupling Proton Translocation to ATP Synthesis

Proton translocation causes a rotation of the Fo unit and the central shaft g subunit This causes a conformational change within all the three ab pairs The conformational change in one of the three pairs promotes condensation of ADP and Pi into ATP The biggest conformation change allows ATP to leave

What are the products of pyruvate dehydrogenase?

Pyruvate dehydrogenase produces acetyl-CoA , NADH , and CO2 from the oxidative decarboxylation of pyruvate.

anaplerotic reactions

Reactions that replenish depleted tricarboxylic acid cycle intermediates Intermediates in the citric acid cycle can be used in biosynthetic pathways (removed from cycle) Must replenish the intermediates in order for the cycle and central metabolic pathway to continue 4-carbon intermediates are formed by carboxylation of 3-carbon precursors

oxidation of molecules

Reduced organic compounds serve as fuels from which electrons can be stripped off during oxidation The oxidation levels of carbon in biomolecules Each compound is formed by oxidation of the red carbon in the compound shown immediately above Carbon dioxide is the most highly oxidized form of carbon found in living systems.

Free Energy of Electron Transport

Reduction Potential (E) deltaE' = E′(e- acceptor) - E′(e- donor) deltaG′ = -nFdeltaE' For negative deltaG need positive deltaE E(acceptor) > E(donor) Electrons are spontaneously transferred from molecules with lower (more negative) to higher (more positive) reduction potential. Free Energy released is used to pump protons, storing this energy as the electrochemical gradient

reduction potential

Reduction potential (E) -Affinity for electrons- higher E, higher affinity -Electrons transferred from molecules with lower E to molecules with higher E -∆E°' assume a pH of 7 and concentrations of 1M ∆E°' = E°'(e- acceptor) - E°'(e- donor) acceptor becomes reduced DE°' = (RT/nF)ln(Keq) = - DG°'/nF ∆G°' = -nF∆E°' For negative DG°' need positive DE°' E(acceptor) > E(donor) R is the gas constant (0.00832 kJ/mol·K) F is the Faraday constant (96.5 kJ/V·mol) T is the absolute temperature (in Kelvin) n is the number of electrons transferred

Regulation of the Citric Acid Cycle

Regulation occurs at Steps 1, 2, 4, and 5. High energy molecules (ATP, Acetyl-CoA, NADH) inhibit while low-energy molecules (ADP, AMP, CoA, NAD+) activate these steps Regulated at highly thermodynamically favorable and irreversible steps - Pyruvate dehydrogenase, citrate synthase, isocitrate dehydrogenase and α-ketoglutarate dehydrogenase General regulatory mechanism - Activated by substrate availability - Inhibited by product accumulation - Overall products of the pathway are NADH and ATP Inhibitors: NADH and ATP Activators: NAD+ and AMP or ADP Enzymes may be associated to allow substrate channeling Regulation of metabolite flow from the PDH complex through the citric acid cycle in mammals. The PDH complex is allosterically inhibited when [ATP]/[ADP], [NADH]/[NAD+], and [acetyl-CoA]/[CoA] ratios are high, indicating an energy-sufficient metabolic state. When these ratios decrease, allosteric activation of pyruvate oxidation results. The rate of flow through the citric acid cycle can be limited by the availability of the citrate synthase substrates, oxaloacetate and acetyl-CoA, or of NAD+, which is depleted by its conversion to NADH, slowing the three NAD-dependent oxidation steps. Feedback inhibition by succinyl-CoA, citrate, and ATP also slows the cycle by inhibiting early steps. In muscle tissue, Ca2+ signals contraction and, as shown here, stimulates energy-yielding metabolism to replace the ATP consumed by contraction.

regulation of cellular respiration

Regulation of the ATP-producing pathways. This diagram shows the interlocking regulation of glycolysis, pyruvate oxidation, the citric acid cycle, and oxidative phosphorylation by the relative concentrations of ATP, ADP, and AMP, and by NADH. High [ATP] (or low [ADP] and [AMP]) produces low rates of glycolysis, pyruvate oxidation, acetate oxidation via the citric acid cycle, and oxidative phosphorylation. All four pathways are accelerated when the use of ATP and the formation of ADP, AMP, and Pi increase. The interlocking of glycolysis and the citric acid cycle by citrate, which inhibits glycolysis, supplements the action of the adenine nucleotide system. In addition, increased levels of NADH and acetyl-CoA also inhibit the oxidation of pyruvate to acetyl-CoA, and a high [NADH]/[NAD+] ratio inhibits the dehydrogenase reactions of the citric acid cycle (see Fig. 16-19).

NADPH regulates partitioning into glycolysis vs. pentose phosphate pathway

Role of NADPH in regulating the partitioning of glucose 6-phosphate between glycolysis and the pentose phosphate pathway When NADPH is forming faster than it is being used for biosynthesis and glutathione reduction [NADPH] rises and inhibits the first enzyme in the pentose phosphate pathway As a result, more glucose 6-phosphate is available for glycolysis

citric acid cycle intermediates are amphibolic

Role of the citric acid cycle in anabolism. Intermediates of the citric acid cycle are drawn off as precursors in many biosynthetic pathways. Shown in red are four anaplerotic reactions that replenish depleted cycle intermediates

citric acid cycle step 5 generation of GTP through Thioester bond cleavage

Some isozymes use ADP and make ATP (brain, heart, muscle) GTP form predominates in liver and kidney

lipid anchors

Some membrane proteins are lipoproteins. They contain a covalently linked lipid molecule. - long-chain fatty acids - isoprenoids - sterols - glycosylated phosphatidylinositol ( GPI) The lipid can become part of the membrane. The protein is now anchored to the membrane. - reversible process if enzyme can cleave lipid moiety off of protein - allows targeting of proteins - Some, such as GPI anchors are found only on the outer face of plasma membrane.

integral membrane proteins ( transmembrane proteins)

Span the entire membrane or linked to membrane by lipid moiety Have asymmetry relative to the membrane - Different segments in different compartments Tightly associated with membrane - Hydrophobic stretches in the protein interact with the hydrophobic regions of the membrane Removed by detergents that disrupt the membrane Purified integral membrane proteins still have phospholipids associated with them Bacteriorhodopsin, a membrane-spanning protein. (PDB ID 2AT9) The single polypeptide chain folds into seven hydrophobic α helices, each of which traverses the lipid bilayer roughly perpendicular to the plane of the membrane. The seven transmembrane helices are clustered, and the space around and between them is filled with the acyl chains of membrane lipids. The light-absorbing pigment retinal (see Fig. 10-21) is buried deep in the membrane in contact with several of the helical segments (not shown). The helices are colored to correspond with the hydropathy plot in Figure 11-12b.

Membrane Dynamics: Transverse Diffusion

Spontaneous flips from one leaflet to another are rare because the charged head group must transverse the hydrophobic tail region of the membrane. uncatalyzed transbilayer flip-flip diffusion very slow Motion of single phospholipids in a bilayer. (a) Uncatalyzed movement from one leaflet to the other is very slow

Stages of Fatty Acid Oxidation

Stage 1 consists of oxidative conversion of two-carbon units into acetyl-CoA via β-oxidation with generation of NADH and FADH2 - involves oxidation of β carbon to thioester of fatty acyl-CoA Stage 2 involves oxidation of acetyl-CoA into CO2 via citric acid cycle with generation NADH and FADH2 Stage 3 generates ATP from NADH and FADH2 via oxidative phosphorylation Ex. palmitate 16:0 Compare this to the oxidation of glucose 4 e- from glycolysis 20 e- from pyruvate through the citric acid cycle Or 4e-/carbon Fatty acid oxidation yields 5.75 e-/carbon (due to starting at more reduced state) (2e- can be in form of FADH2 or NADH) Stages of fatty acid oxidation. Stage 1: A long-chain fatty acid is oxidized to yield acetyl residues in the form of acetyl-CoA. This process is called β oxidation. Stage 2: The acetyl groups are oxidized to CO2 via the citric acid cycle. Stage 3: Electrons derived from the oxidations of stages 1 and 2 pass to O2 via the mitochondrial respiratory chain, providing the energy for ATP synthesis by oxidative phosphorylation.

Why does starting with NADH produce more ATP than succinate/FADH2?

Starting with NADH produces more ATP compared to FADH2 because Complex I pumps four protons into the IMS for every NADH that is oxidized Complex II does not pump any protons.

sequence of events in the citric acid cycle

Step 1: C-C bond formation to make citrate Step 2: Isomerization via dehydration/rehydration Steps 3-4: Oxidative decarboxylations to give 2 NADH Step 5: Substrate-level phosphorylation to give GTP Step 6: Dehydrogenation to give reduced FADH2 Step 7: Hydration Step 8: Dehydrogenation to give NADH

fumarase reaction

Stereospecific - Addition of water is always trans and forms L-malate - OH- adds to fumarate... then H+ adds to the carbanion - Cannot distinguish between inner carbons, so either can gain -OH Slightly thermodynamically favorable/reversible - Product concentration kept low to pull reaction forward

induced fit in the citrate synthase

Structure of citrate synthase. The flexible domain of each subunit undergoes a large conformational change on binding oxaloacetate, creating a binding site for acetyl-CoA. (a) Open form of the enzyme alone ( PDB ID 5CSC); (b) closed form with bound oxaloacetate and a stable analog of acetyl-CoA ( carboxymethyl-CoA) (derived from PDB ID 5CTS). In these representations one subunit is colored tan and one green.

What enzyme of Citric Acid Cycle is not found in the mitochondrial matrix? Where is it located? Why does it have to be located there?

Succinate dehydrogenase , is in the inner membrane due to the fact that this enzyme is the part of the complex II it is part of Complex II is that it utilizes FAD/FADH2 for its redox reaction which is a prosthetic group To get the electrons from FADH2 into the electron transport chain, succinate dehydrogenase is in the inner membrane as part of Complex II fact: (The other evolutionary alternative is what happens with acyl-CoA dehdrogenase which passes the electrons from FADH2 to the Electron Transferring Flavoprotein which moves them to the membrane where it passes them off to the ETF:Q oxidoreductase which redues ubiquinone to ubiquinol.)

Succinyl-CoA Synthetase Reaction

SuccinylCoA + GDP + Pi <--> Succinate + GTP + HSCoA Substrate level phosphorylation Energy of breaking thioester bond allows for incorporation of inorganic phosphate to make GTP - Phosphate displaces CoA to make succinyl phosphate - Phosphate transferred to make phospho-enzyme intermediate-succinate leaves - Phosphate added to GDP to produce GTP, which can be converted to ATP Slightly thermodynamically favorable/reversible Product concentration kept low to pull forward

Summary of the electron flow in the respiratory chain

Summary of the flow of electrons and protons through the four complexes of the respiratory chain. Electrons reach Q through Complexes I and II. The reduced Q (QH2) serves as a mobile carrier of electrons and protons. It passes electrons to Complex III, which passes them to another mobile connecting link, cytochrome c. Complex IV then transfers electrons from reduced cytochrome c to O2. Electron flow through Complexes I, III, and IV is accompanied by proton flow from the matrix to the intermembrane space. Recall that electrons from β oxidation of fatty acids can also enter the respiratory chain through Q The structures shown here are from several sources: Complex I, Thermus thermophilus (PDB ID 3M9S); Complex II, porcine heart (PDB ID 1ZOY); Complex III, bovine heart (PDB ID 1BGY); cytochrome c, equine heart (PDB ID 1HRC); Complex IV, bovine heart (PDB ID 1OCC).

Summary of the electron flow in the respiratory chain

Summary of the flow of electrons and protons through the four complexes of the respiratory chain. Electrons reach Q through Complexes I and II. The reduced Q (QH2) serves as a mobile carrier of electrons and protons. It passes electrons to Complex III, which passes them to another mobile connecting link, cytochrome c. Complex IV then transfers electrons from reduced cytochrome c to O2. Electron flow through Complexes I, III, and IV is accompanied by proton flow from the matrix to the intermembrane space. Recall that electrons from β oxidation of fatty acids can also enter the respiratory chain through Q (see Fig. 19-8). The structures shown here are from several sources: Complex I, Thermus thermophilus (PDB ID 3M9S); Complex II, porcine heart (PDB ID 1ZOY); Complex III, bovine heart (PDB ID 1BGY); cytochrome c, equine heart (PDB ID 1HRC); Complex IV, bovine heart (PDB ID 1OCC).

types of transport

Summary of transporter types. Some types (ionophores, ion channels, and passive transporters) simply speed transmembrane movement of solutes down their electrochemical gradients, whereas others (active transporters) can pump solutes against a gradient, using ATP or a gradient of a second solute to provide the energy.

oxaloacetate to phosphoenolpyruvate ( PEP )

Synthesis of phosphoenolpyruvate from pyruvate. (b) In the cytosol, oxaloacetate is converted to phosphoenolpyruvate by PEP carboxykinase The CO2 incorporated in the pyruvate carboxylase reaction is lost here as CO2 The decarboxylation leads to a rearrangement of electrons that facilitates attack of the carbonyl oxygen of the pyruvate moiety on the γ phosphate of GTP

bicarbonate transporter is an antiporter

System to transport CO2 to lungs. Antiport speeds bicarbonate transport and maintains the electrochemical potential across the membrane. In tissues: CO2 diffuses in and bicarbonate is transported out In lungs: bicarbonate transported in and CO2 diffuses out Chloride-bicarbonate exchanger of the erythrocyte membrane. This cotransport system allows the entry and exit of HCO3- without changing the membrane potential. Its role is to increase the CO2-carrying capacity of the blood. The top half of the figure illustrates the events that take place in respiring tissues; the bottom half, the events in the lungs.

ethanol fermentation

TPP and Mg2+ are cofactors

NADH and FADH2 help make ATP

Takes the equivalent of 2 ATP to do initial activation to palmitoyl-CoA so net gain is 106 ATP. Activation expense the same for all lengths of fatty acids

Why does gluconeogenesis only bypass 3 reactions of glycolysis?

The 3 reactions are irreversible in glycolysis and must be bypass in gluconeogenesis, because this reactions are thermodynamically favorable and regulated, also different enzymes must used in different pathways. Three reactions in glycolysis are thermodynamically favorable in that direction and irreversible so they must use a different route and enzymes to go in the opposite direction in gluconeogenesis.

ion channels provide passive transport down the concentration gradient

The K+ channel of Streptomyces lividans. (PDB ID 1BL8) (a) Viewed in the plane of the membrane, the channel consists of eight transmembrane helices (two from each of four identical subunits), forming a cone with its wide end toward the extracellular space. The inner helices of the cone (lighter colored) line the transmembrane channel, and the outer helices interact with the lipid bilayer. Short segments of each subunit converge in the open end of the cone to make a selectivity filter. (b) This view, perpendicular to the plane of the membrane, shows the four subunits arranged around a central channel just wide enough for a single K+ ion to pass. (c) Diagram of a K+ channel in cross section, showing the structural features critical to function. Carbonyl oxygens (red) of the peptide backbone in the selectivity filter protrude into the channel, interacting with and stabilizing a K+ ion passing through. These ligands are perfectly positioned to interact with each of four K+ ions but not with the smaller Na+ ions. This preferential interaction with K+ is the basis for the ion selectivity.

actual deltaG of ATP hydrolysis to ADP differs from deltaG'

The actual free-energy change in a process depends on: -The standard free energy (-30.5 kJ/mol for hydrolysis of ATP) -The actual concentrations of reactants and products The free-energy change is more favorable if the ratio of reactant's concentration to product's concentration exceeds that at standard concentration -[ATP] kept high in cells If [ATP] levels get low, fewer molecules, but also ΔG becomes less negative-less energy to drive reactions deltaG = deltaG' + RTln [ADP]-[Pi] / [ATP]

Fats Provide Efficient Fuel Storage

The advantage of fats over polysaccharides: - Fatty acids carry more energy per carbon because they are more reduced - Fatty acids carry less water per gram because they are nonpolar • Glucose and glycogen are for short-term energy needs, quick delivery • Fats are for long-term (months) energy needs, good storage, slow delivery

each round produces an acetyl-CoA and shortens the chain by 2 carbons

The b-oxidation pathway. (b) Six more passes through the pathway yield seven more molecules of acetyl-CoA, the seventh arising from the last two carbon atoms of the 16-carbon chain. Eight molecules of acetyl-CoA are formed in all.

In the proton motive force, there is the chemical component and the electrical component. What are each of these components?

The chemical component of the proton motive force is the difference between the concentration of protons on the P side compared to the N side. The P side has a much larger concentration of protons. The electrical component is the difference in charge between the P and N sides. The P side is positively charged and the N side is negatively charged. Therefore, the electrochemical gradient makes it very favorable for protons to leave the P side and enter the N side.

chemical logic of glycolysis

The chemical logic of the glycolytic pathway. In this simplified version of the pathway, each molecule is shown in a linear form, with carbon and hydrogen atoms not depicted, in order to highlight chemical transformations. Remember that glucose and fructose are present mostly in their cyclized forms in solution, although they are transiently present in linear form at the active sites of some of the enzymes in this pathway. The preparatory phase, steps 1 to 5, converts the six-carbon glucose into two three-carbon units, each of them phosphorylated. Oxidation of the three-carbon units is initiated in the payoff phase. To produce pyruvate, the chemical steps must occur in the order shown.

The Citric Acid Cycle is considered to be amphibolic. What does this mean?

The citric acid cycle is considered amphibolic because the intermediates can be used in anabolic and catabolic pathways.

What is the mobile electron carrier that moves between Complex I and Complex III OR Complex II to Complex III? (remember electrons are not moved between Complexes I and II.)

The coenzyme Q or ubiquinol QH2.

Name the electron carriers that are in Complexes I. How many electrons do they each carry?

The electron carriers in complex I and the number of protons each carrier can take. are flavin mononucleotide FMN = takes 2 iron sulfur centers = takes 1 coenzyme Q ( ubiquinone ) = takes 2 Note: that FMN can pick up two electrons from NADH and pass one at a time to the iron sulfur cluster. Ubiquinone can take one at a time until it has the full two it needs.

What are the products of the extra reactions for ethanol fermentation? How many reactions are involved?

The final products of ethanol fermentation are CO2 , ethanol, and NAD+ The reduction of pyruvate to ethanol through fermentation takes place in 2 steps.

For active transport, what are the two potential energy sources used to transport molecules?

The first is the ability of protein domains to hydrolyse ATP and gain energy in order to perform the active transport, the other one is the use of the ions and thier movement down their electrochemical gardient, which also porvide an energy in oreder to perform the movent of the other molecule againts it's concentration gradient, as an example the Na+ - glucose symporter.

a. How many redox reactions occur in the pyruvate dehydrogenase complex? b. What molecule is reduced during each reaction?

The first one is the oxidation of aldehyde to thioester, FAD to FADH while lipoyl cofactor is reoxidized, NAD+ to NADH while FADH2 is oxidized, total of three redox reactions. Lipoyl cofactor, FAD, NAD+

Why is the 3rd step that produces Fructose 1,6 bis phosphate called the first committed step of glycolysis?

The formation of fructose 1,6-bisphosphate is an irreversible reaction, it highly regulated by ATP or other metabolites and it can not be utilized by other pathways such as glyconeogenesis or pentose phosphate pathway, thus, the only way it can be further breakdown is through energetically silly glycolysis. Fructose 1,6 bisphosphate can only be used in glycolysis. Previous products, glucose-6P and fructose-6P can be utilized by other pathways.

What is the free energy that is produced by the favorable transfer of electrons in the electron transport chain used for?

The free energy is used to transport protons to the IMS , against their electrochemical gradient.

Why make 1,3 bisphosphoglycerate and phosphoenolpyruvate?

The idea behind the formation of 1,3 bisphosphoglycerate (BPG) and the phosphoenolpyruvate are their highly negative free energy for the hydrolysis of the phosphate group around -70kJ/mole, that energy can be easily used for the formation of the main energy currency of the cell.

Under what conditions is the non-oxidative phase used?

The idea of the non-oxidative phase is to regenerate glucose 6P from ribose 5P, and then use it to produce more NADPH that the cell required more than ribose 5P It's mostly used in tissues and organs with high demand for NADPH such as adipose tissue and liver. So the non-oxidative phase is used when NADPH is needed but ribose-5phosphate is not needed

How can membrane structure be different between organisms or between different organelles?

The membrane is dynamic, due to its difference in composition, such as ratio of lipids to proteins, types of phospholipids and types of sterols present are varies. As well as organisms and it's conditions and habitats, thus membrane must be adjusted to certain conditions in order to maintain its proper fluidity for proper functioning.

How are the substrates and products of ATP synthase moved across the inner mitochondrial membrane? (name the transporter, what is transported and the direction of transport)

The phosphate (substrate) and H+ moves into the matrix from intermembrane space through phosphate translocase ( symporter ), and the ADP (substrate moves into the matrix) and ATP (product moves out of the matrix) through adenine nucleotide translocase ( antiporter )

What is the point of glycolysis?

The point of glycolysis is to generate energy in the form of ATP and NADH by oxidizing glucose. Glycolysis also forms pyruvate, which is a precursor for other metabolic pathways.

What is the product of glycogen phosphorylase?

The product of glycogen phosphorylase is glucose 1-phosphate.

oxidative phosphorylation

The production of ATP using energy derived from the redox reactions of an electron transport chain; the third major stage of cellular respiration. final stage of cellular respiration

Chemiosmotic energy coupling requires membranes

The proton gradient needed for ATP synthesis can be stably established across a membrane that is impermeable to ions - Plasma membrane in bacteria - Inner membrane in mitochondria - Thylakoid membrane in chloroplasts Membrane must contain proteins that couple the "downhill" flow of electrons in the electron-transport chain with the "uphill" flow of protons across the membrane Membrane must also contain a protein that couples the "downhill" flow of protons to the phosphorylation of ADP

Cytochrome c

The second mobile electron carrier A soluble heme-containing protein in the intermembrane space Heme iron can be either Fe3+ (oxidized) or Fe2+ (reduced) Cytochrome c carries a single electron from complex III to complex IV Prosthetic groups of cytochromes. (a) Each group consists of four five-membered, nitrogen-containing rings in a cyclic structure called a porphyrin. The four nitrogen atoms are coordinated with a central Fe ion, either Fe2+ or Fe3+. Iron protoporphyrin IX is found in b-type cytochromes and in hemoglobin and myoglobin (see Fig. 4-17). Heme c is covalently bound to the protein of cytochrome c through thioether bonds to two Cys residues. Heme a, found in a-type cytochromes, has a long isoprenoid tail attached to one of the five-membered rings. The conjugated double-bond system (shaded light red) of the porphyrin ring has delocalized π electrons that are relatively easily excited by photons with the wavelengths of visible light, which accounts for the strong absorption by hemes (and related compounds) in the visible region of the spectrum.

What is the signal that glucose-6P should go into the pentose phosphate pathway?

The signal that glucose-6P should go to pentose phosphate is that NADPH is low

How is the structure of ABC transporters designed for active transport?

The transmembrane portion is responsible for moving the molecule to be transported across the membrane while the ATP binding domains hydrolyze the ATP to provide the energy for transport. ABC transporter's structure has transmembrane domains (outside) and nucleotide binding domains facing the cytoplasm with alpha helices that span the membrane (6 each). ATP hydrolysis causes the protein configuration to change which allows transport molecules against the concentration gradient so the structure helps active transport.

Which secondary structure is most commonly found in membrane proteins?

The transmembrane segment of integral membrane proteins are predominantly composed of alpha helices.

What are the two phases of the pentose phosphate pathway and what are their main products?

The two phases are the oxidative phase and the non-oxidative phase. The products of the oxidative phase are NADPH and Ribose 5-Phosphate The product of the non-oxidative phase is Glucose 6-Phosphate. Extra info: Ribose-5phosphate is the precursor for nucleic acids.

glycolysis : the payoff phase

The two phases of glycolysis. For each molecule of glucose that passes through the preparatory phase (a), two molecules of glyceraldehyde 3-phosphate are formed; both pass through the payoff phase (b). Pyruvate is the end product of the second phase of glycolysis. For each glucose molecule, two ATP are consumed in the preparatory phase and four ATP are produced in the payoff phase, giving a net yield of two ATP per molecule of glucose converted to pyruvate. The numbered reaction steps correspond to the numbered headings in the text discussion. Keep in mind that each phosphoryl group, represented here as P , has two negative charges (—PO32-).

glycolysis : the preparatory phase

The two phases of glycolysis. For each molecule of glucose that passes through the preparatory phase (a), two molecules of glyceraldehyde 3-phosphate are formed.

Name the two types of proteins associated with lipids. Which type requires the destruction of the membrane to lose association?

The two types of proteins associated with lipids are peripheral membrane proteins and integral membrane proteins. Integral membrane proteins require the destruction of the membrane to dissociate.

How many protons are pumped (moved by a separate transport process) In Complex I? In Complex II?

There are 4 protons H+ pumped in complex I , and complex II does not pumps protons , only I and IV has subunits to do pump or pumbing protons.

Facets of pyruvate

Three possible catabolic fates of the pyruvate formed in glycolysis Pyruvate also serves as a precursor in many anabolic reactions, not shown here.

Most Common Redox Reactions Continued

Transfer of electrons to molecular oxygen, which is reduced to water or H2O2 during this process. - Catalyzed by oxidases Incorporation of one or both oxygen atoms from O2 into a substrate. - Catalyzed by oxygenases

The most common types of redox reactions in biological systems

Transfer of single electrons with or without simultaneous transfer of protons Enzymes involved in this type of redox reaction typically require cofactors - hemes (change in oxidation state of iron between Fe2+ and Fe3+) - iron-sulfur proteins - copper ions (Cu+/Cu2+) - flavin nucleotides (FMN or FAD) Occur predominantly in mitochondria as part of the electron transfer chain, and in chloroplasts or cyanobacteria as part of the light reactions in photosynthesis. Transfer of a hydride ion (one proton plus two electrons). - NAD+/NADH and NADP+/NADPH are usually involved in these reactions - Catalyzed by dehydrogenases or reductases

Complex I is a proton pump

Transfer of two electrons from NADH to ubiquinone is accompanied by a transfer of protons from the matrix (N) to the intermembrane space (P) Experiments suggest that about four protons are transported per one NADH NADH + Q + 5H+N → NAD+ + QH2 + 4 H+P Reduced coenzyme Q picks up two protons Protons are transported by subunits that have structural similarity to transporters, probably at the same time

Amino acids in membrane proteins cluster in distinct regions

Transmembrane segments are predominantly hydrophobic Tyr and Trp cluster at nonpolar/polar interface Charged amino acids are only found in aqueous domains Tyr and Trp residues of membrane proteins clustering at the water-lipid interface. The detailed structures of these five integral membrane proteins are known from crystallographic studies. The K+ channel (PDB ID 1BL8) is from the bacterium Streptomyces lividans (see Fig. 11-47); maltoporin (PDB ID 1AF6), outer membrane phospholipase A ( OmpLA, PDB ID 1QD5), OmpX (PDB ID 1QJ9), and phosphoporin E (PDB ID 1PHO) are proteins of the outer membrane of E. coli. Residues of Tyr and Trp are found predominantly where the nonpolar region of acyl chains meets the polar head group region. Charged residues ( Lys, Arg, Glu, Asp) are found almost exclusively in the aqueous phases.

2 types of active transport

Two types of active transport. (a) In primary active transport, the energy released by ATP hydrolysis drives solute (S1) movement against an electrochemical gradient. (b) In secondary active transport, a gradient of ion X (S1) (often Na+) has been established by primary active transport. Movement of X (S1) down its electrochemical gradient now provides the energy to drive cotransport of a second solute (S2) against its electrochemical gradient. primary active transport - ADP + Pi, ATP, S1 secondary active transport - ADP + pi, ATP, S1, S2

What is the substrate for glycogen synthesis?

UDP-Glucose is the substrate for glycogen synthesis.

What are the products of UDP glucose:galactose-1phosphate uridylyltransferase?

UDP-galactose and glucose-1P

Coenzyme Q or Ubiquinone

Ubiquinone (Q, or coenzyme Q). Complete reduction of ubiquinone requires two electrons and two protons, and occurs in two steps through the semiquinone radical intermediate.

Coenzyme Q aka Ubiquinone

Ubiquinone is a lipid-soluble conjugated dicarbonyl compound that readily accepts electrons Can transfer electrons 1 or 2 at a time - Upon accepting an electron, it picks up a proton - Full reduction produces an alcohol, ubiquinol Ubiquinol can freely diffuse in the membrane, carrying electrons with protons Ubiquinol is a mobile electron carrier transporting electrons from Complexes I or II to Complex III Ubiquinone is referred to in future slides as Q and ubiquinol is QH2

flavin cofactors allow single and double electron transfers

Used in oxidative phosphorylation and photosynthesis as an electron carrier prosthetic groups-tightly bound to enzymes Flavin Adenine dinucleotide (FAD) or Flavin mononucleotide can accept one electron and one hydrogen at a time to finally make FADH2 or FMNH2 FADH2 or FMNH2 can also pass one electron at a time Can also pass or accept 2 electrons and 2 hydrogens-usually to NAD+ or from NADH Oxidized and reduced FAD and FMN. FMN consists of the structure above the dashed line on the FAD (oxidized form). The flavin nucleotides accept two hydrogen atoms (two electrons and two protons), both of which appear in the flavin ring system. When FAD or FMN accepts only one hydrogen atom, the semiquinone, a stable free radical, forms.

Ubiquinone:cytochrome c Oxidoreductase aka Complex III ubiquinone-cytochrome

Uses two electrons from ubiquinol (QH2) to reduce two molecules of cytochrome c Additionally contains iron-sulfur clusters, cytochrome bs, and cytochrome cs The Q cycle results in four additional protons being added to the IMS Cytochrome bc1 complex (Complex III). (PDB ID 1BGY) The complex is a dimer of identical monomers, each with 11 different subunits. The functional core of each monomer is three subunits: cytochrome b (green) with its two hemes (bH and bL), the Rieske iron-sulfur protein (purple) with its 2Fe-2S centers, and cytochrome c1 (blue) with its heme. This cartoon view of the complex shows how cytochrome c1 and the Rieske iron-sulfur protein project from the P surface and can interact with cytochrome c (not part of the functional complex) in the intermembrane space. The complex has two distinct binding sites for ubiquinone, QN and QP, which correspond to the sites of inhibition by two drugs that block oxidative phosphorylation. Antimycin A, which blocks electron flow from heme bH to Q, binds at QN, close to heme bH on the N (matrix) side of the membrane. Myxothiazol, which prevents electron flow from QH2 to the Rieske iron-sulfur protein, binds at QP, near the 2Fe-2S center and heme bL on the P side. The dimeric structure is essential to the function of Complex III. The interface between monomers forms two caverns, each containing a QP site from one monomer and a QN site from the other. The ubiquinone intermediates move within these sheltered caverns. Complex III crystallizes in two distinct conformations (not shown). In one, the Rieske Fe-S center is close to its electron acceptor, the heme of cytochrome c1, but relatively distant from cytochrome b and the QH2- binding site at which the Rieske Fe-S center receives electrons. In the other, the Fe-S center has moved away from cytochrome c1 and toward cytochrome b. The Rieske protein is thought to oscillate between these two conformations as it is first reduced, then oxidized.

origin of C-atoms in CO2

We have lost 2 CO2 already, so we have a net complete oxidation of glucose after two pyruvates go through the TCA cycle. But its not the actual carbons from pyruvate in each cycle.

Cellular respiration involves the complete oxidation of glucose. What is the molecule that is produced when carbon is fully oxidized? Starting with pyruvate, how many carbons are fully oxidized by the end of the citric acid cycle?

When carbon is fully oxidized, CO2 is produced. Starting with one pyruvate molecule, three carbons are fully oxidized by the end of the citric acid cycle.

When ketone bodies are broken down in the brain, what is produced that can enter an energy producing pathway? What is that pathway?

When ketone bodies are broken down in the brain, acetyl-CoA is produced that can then enter the citric acid cycle and eventually produce ATP through oxidative phosphorylation.

How is the movement of a lipid between leaflets different from moving it within a leaflet (transverse vs lateral movement)?

When lipids are within a leaflet they undergo very fast lateral movement. When lipids are moved between the leaflets (transverse movement), movement is very slow because the hydrophilic head group has to pass through the hydrophobic section of the membrane which is not favorable. In order to overcome the unfavorable transverse movement, transporters move lipids between the leaflets.

the citric acid cycle ( CAC )

[carbohydrate metabolism] takes place in the mitochondrial matrix; oxidizes acetyl-coA into co2 and generates nadh and fadh2 and gtp; yields 6 nadh, 2 fadh2 and 2 gtp / glucose molecule) Reactions of the citric acid cycle. The carbon atoms shaded in pink are those derived from the acetate of acetyl-CoA in the first turn of the cycle; these are not the carbons released as CO2 in the first turn. Note that in succinate and fumarate, the two-carbon group derived from acetate can no longer be specifically denoted; because succinate and fumarate are symmetric molecules, C-1 and C-2 are indistinguishable from C-4 and C-3. The red arrows show where energy is conserved by electron transfer to FAD or NAD+, forming FADH2 or NADH + H+. Steps 1, 3, and 4 are essentially irreversible in the cell; all other steps are reversible. The nucleoside triphosphate product of step 5 may be either ATP or GTP, depending on which succinyl-CoA synthetase isozyme is the catalyst.

alpha-ketoglutarate a-ketoglutarate dehydrogenase reaction

a compound that participates in the formation of nonessential amino acids during transamination Catalyzes last oxidative decarboxylation - Net full oxidation of all carbons of acetyl-CoA After two turns of the cycle (2 pyruvates from payoff phase of glycolysis)- complete oxidation of all carbons from glucose Carbons not directly from glucose because carbons came from oxaloacetate, not from acetyl-CoA Ketone on α-ketoglutarate oxidized to a thioester NAD+ is reduced to NADH Succinyl-CoA has a higher-energy thioester bond Highly thermodynamically favorable/irreversible - Regulated by product inhibition

metabolism = is the sum of the chemical reactions in an organism catabolism = is the energy releasing processes anabolism = is the energy using processes

a metabolic pathway is a sequence of enzymatically catalyzed chemical reactions in a cell metabolic pathway is a sequence of enzymatically catalyzed chemical reactions in a cell metabolic pathways are determined by enzymes enzymes are encoded by genes

proton motive force ( PMF)

a source of energy resulting from the separation of protons from hydroxyl ions across the cytoplasmic membrane, generating a membrane potential The proteins in the electron-transport chain created the electrochemical proton gradient by one of three means: - Actively transporting protons across the membrane (aka pumping of protons) -- Complex I and Complex IV - Chemically removing protons from the matrix -- Reduction of ubiquinone (Q) (complex III) and reduction of oxygen (complex IV) - Release of protons into the intermembrane space -- Oxidation of 2 ubiquinol ( QH2) Proton-motive force. The inner mitochondrial membrane separates two compartments of different [H+], resulting in differences in chemical concentration (∆pH) and charge distribution (∆ψc) across the membrane. The net effect is the proton-motive force (∆G), which can be calculated as shown here. This is explained more fully in the text. In actively respiring mitochondria ΔpH=0.75 (intermembrane space has lower pH) Δψ= 0.15 to 0.2V

By the end of the citric acid cycle, how many NADH and FADH2 are produced a) starting with 1 acetyl CoA b) starting with 1 pruvate c) starting with glucose including glycolysis

a) 3 NADH and 1 FADH2 b) 4 NADH and 1 FADH2 c) 10 NADH and 2 FADH2 (explanation for c:) 2 NADH are formed during glycolysis and then pyruvate dehydrogenase and the citric acid cycle are run twice (once for each of 2 pyruvates from glycolysis) so 8 NADH and 2 FADH2

The prepatory phase uses 2 ATP. a) Name the 2 molecules that are phosphorylated. b) Where are the phosphates at the end of the prepatory phase?

a) Glucose is phosphorylated to form glucose 6-phosphate, and fructose 6-phosphate is phosphorylated to form fructose 1,6-bisphosphate b) At the end of the prepatory phase, there is 1 phosphate group on each of the 2 glyceraldehyde 3-phosphate molecules produced

a. Under what conditions is the Inhibitor of F1 needed? b. What does the Inhibitor of F1 prevent?

a. F1 is needed under low oxygen/low pH b. it prevents ATPase from turning backwards

a. Name the 2 shuttles that move the e- from NADH from glycolysis into the electron transport chain. b. What molecule binds to a complex in electron transport chain for each shuttle? What complex does it bind to? c. How many ATP are made as a result of each shuttle?

a. malate-aspartate shuttle and glycerol-3P shuttle b. Malate-aspartate shuttle- NADH binds to Complex I Glycerol-3P shuttle- ubiquinol binds to Complex III c. The values given are for two NADH coming from glycolysis so it's 2.5 ATP per NADH for the malate-aspartate shuttle and 1.5 ATP per NADH for the glycerol-3P shuttle

a. What happens with each of the conformation of ATP synthase to synthesize ATP? βADP, βATP, βempty b. What causes the change between the confromations?

a. ßATP forms ATP from ADP and Pi ßempty releases the ATP ßADP binds ADP and Pi b. specifically the γ (gamma) shaft rotates to cause a change in the conformation of the subunits in F1

a. How many NADH and FADH2 are produced for each cycle of β-oxidation? b. Where do the electrons from NADH and FADH2 enter the electron transport chain (which complex or mobile electron carrier)? c. Which of the two molecules requires the electron transferring flavoprotein to transport electrons?

a. 1 FADH2 and 1 NADH. b. The electrons from FADH2 enter electron transport at ubiquinol. The electron transferring flavoprotein is not part of the electron transport chain proper. c. FADH2 requires the ETF

What is the ΔG of transporting Na+ out of a cell when [Na+] inside is 12μM and [Na+] outside is 145 μM and the membrane potential is 0.05V (inside negative) at 310K? deltaG = RTln(c2/c1) + zFdeltaY ΔGt=RTln(C2/C1) + zFΔΨ R=0.0083 kJ/mol F= 96.5 kJ/molV Remember when answering questions that some of these values require a correct sign as well as a value. a) What are C2 and C1? b) What is z? c) What is ΔΨ? d) What is ΔGt e) Can the Na+ be transported out of the cell passively or must active transport be used?

a. C2-[outside], C1-[inside] b. +1 c. + 0.05V (positive because the Na ions are being moved to the positive side of the membrane) d. using the ΔΨ of +0.05V in the second term would give a value of 11.2 kJ/mol which is still not favorable. e. Active transport must be used, which required ATP, in order to move sodium ions against its [gradient].

a. What molecule from the break down of glycereol enters glycolysis? (I want the first molecule that qualifies) b. This is a two step process. What is the main product of the first step?

a. Dihydroxyacetone phosphate enters glycolysis from the breakdown of glycerol. b. The main product of the first step is glycerol 3-phosphate.

a. What do the isomerase and the reductase do as part of breaking down unsaturated fatty acids? b. Which one(s) are needed for double bonds that start at odd numbered carbons? even numbered carbons?

a. Isomerase converts double bonds starting at carbon 3 to trans double bonds starting at carbon 2 Reductase reduces double bonds that do not start at carbon 3 b. Isomerase is needed for fatty acids that have a double bond starting at an odd numbered carbon. Both isomerase and reductase are needed for fatty acids that have a double bond starting at an even numbered carbon.

a. Why does the Citric Acid Cycle need anapleurotic reactions? b. What 4 carbon molecules are the products of these reactions?

a. It needs to replace intermediates that go to other biosythetic pathways. Without the replenished intermediates oxaloacetate can quickly deplete and the cycle can not be run. b. malate and oxaloacetate are made

a. Where does the energy come from to make GTP? b. What enzyme catalyzes this reaction?

a. The energy to form GTP comes from the cleavage of the thioester bond within Succinyl-CoA. b. Succinyl-CoA synthetase catalyzes the reaction. Note: Synthetases involve a nucleotide, in this case GTP. Synthases do not include a nucleotide.

a. What enzyme catalyzes the first step of β-oxidation of fatty acids in peroxisomes? b. What happens to the FADH2 that is produced in this reaction?

a. The first step of ß-oxidation beta-oxidation in peroxisomes is catalyzed by acyl-CoA oxidase. b. FADH2 passes its electrons to molecular oxygen, creating hydrogen peroxide

a. What are the two functional units of ATP synthase? b. What does each functional units do?

a. The two functional subunits of ATP synthase are F0 and F1. b. F0 transports protons from the IMS to the matrix, transferring energy to F1. F1 catalyzes the synthesis of ATP from ADP and Pi.

What is the name of the transporter of long chain fatty acids in the mitochondrial inner membrane?

acyl-carnitine/carnitine transporter.

formation of ketone bodies

aka acyl CoA acetyltransferase Formation of ketone bodies from acetyl-CoA. Healthy, well-nourished individuals produce ketone bodies at a relatively low rate. When acetyl-CoA accumulates (as in starvation or untreated diabetes, for example), thiolase catalyzes the condensation of two acetyl-CoA molecules to acetoacetyl-CoA, the parent compound of the three ketone bodies. The reactions of ketone body formation occur in the matrix of liver mitochondria. The six-carbon compound β-hydroxy-β-methylglutaryl-CoA (HMG-CoA) is also an intermediate of sterol biosynthesis, but the enzyme that forms HMG-CoA in that pathway is cytosolic. HMG-CoA lyase is present only in the mitochondrial matrix.

Precursors for gluconeogenesis

animals CAN produce glucose from sugars or proteins - sugars: pyruvate , lactate , or oxaloacetate - protein: from amino acids that can be converted to citric acid cycle intermediates or pyruvate animals can NOT produce glucose from fatty acids through acetyl-CoA and citric acid cycle - product of fatty acid degradation is acetyl-CoA - cannot have a net conversion of acetyl-CoA to oxaloacetate > plants, yeast, and many bacteria can do this by a different pathway, thus producing glucose from fatty acids -major substrates are: 1. Lactate 2. Alanine 3. Glycerol note: Fatty acids are not used

hydrolysis of ATP is highly favorable under standard conditions

better charge separation in products better solvation of products more favorable resonance stabilization of products Chemical basis for the large free-energy change associated with ATP hydrolysis. The charge separation that results from hydrolysis relieves electrostatic repulsion among the four negative charges on ATP. The product inorganic phosphate (Pi) is stabilized by formation of a resonance hybrid, in which each of the four phosphorus-oxygen bonds has the same degree of double-bond character and the hydrogen ion is not permanently associated with any one of the oxygens (Some degree of resonance stabilization also occurs in phosphates involved in ester or anhydride linkages, but fewer resonance forms are possible than for Pi.) A third factor (not shown) that favors ATP hydrolysis is the greater degree of solvation (hydration) of the products Pi and ADP relative to ATP, which further stabilizes the products relative to the reactants.

Step 3: 2nd priming phosphorylation

bis used for 2 phosphates in different places on the molecule di used for 2 phosphates linked together on a molecule rationale - further activation of fructose - allows for 1 phosphate / 3-carbon sugar after step 4 first committed step of glycolysis - fructose 1,6-biphosphate must continue through glycolysis whereas prior products may be utilized by other pathways this process uses the energy of ATP highly thermodynamically favorable / irreversible phosphofructokinase-1 is highly regulated - by ATP, fructose 2,6-bisphosphate and other metabolite - do not burn glucose if there is plenty of ATP

additional bypass reactions

catalyze reverse reaction of opposing irreversible step in glycolysis are irreversible themselves fructose 1,6-biphosphate > fructose 6-phosphate + Pi - enzyme fructose 1,6-biphosphatase - coordinately / oppositely regulated with PFK-1 - deltaG' = -16.3 kJ/mol glucose 6-phosphate > glucose + Pi - enzyme glucose 6-phosphatase - only used in tissues that send glucose out to other parts of the body like liver - deltaG' = -13.8 kJ/mol

What is the product of the rate limiting reaction of the citric acid cycle?

citrate

Naming

citric acid cycle - the first molecule made is citrate - first name for the cycle TCA cycle - tricarboxylic acid cycle - first two molecules made ( citrate and isocitrate) have 3 carboxyl groups Krebs cycle - cycle discovered by Hans Krebs in 1937

disaccharides

each has specific enzyme maltose cleaved by maltase lactose cleaved by lactase sucrose cleaved by sucrase hydrolysis reaction produces monosaccharides

What enzymes catalyze the last two bypass reactions?

fructose-1,6-biphosphatase and glucose-6-phosphatase

galactose

galactokinase synthesizes galactose 1-P from galactose using ATP UDP-glucose galactose 1-phosphate uridylyltransferase trades groups - UDP from UDP-glucose transferred to galactose to C-1 and phosphate from galactose 1-P transferred to glucose at C-1 - results in: UDP-galactose glucose 1-P converted to glucose 6-P by phosphoglucomutase - entrance of sugar into glycolysis UDP-glucose-4-epimerase switches OH on chiral carbon 4 of the galactose portion of UDP-galactose to make UDP-glucose ( galactose is the C4 epimer of glucose )

gluconeogenesis : building carbohydrates

gluconeogenesis can occur in almost all living things long held that mammals cannot convert fatty acids to sugars - may not be true Carbohydrate synthesis from simple precursors The pathway from phosphoenolpyruvate to glucose 6-phosphate is common to the biosynthetic conversion of many different precursors of carbohydrates in animals and plants The path from pyruvate to phosphoenolpyruvate leads through oxaloacetate, an intermediate of the citric acid cycle Any compound that can be converted to either pyruvate or oxaloacetate can therefore serve as starting material for gluconeogenesis This includes alanine and aspartate, which are convertible to pyruvate and oxaloacetate and other amino acids that can also yield three- or four-carbon fragments, the so called glucogenic amino acids Plants and photosynthetic bacteria are uniquely able to convert CO2 to carbohydrates, using the Calvin cycle

only a small amount of energy available in glucose is captured in glycolysis

glucose -- glycolysis deltaG' = -146 kJ/mol --> 2 pyruvate glucose -- full oxidation + 6 O2 deltaG' = -2840 kJ/mol -> 6 CO2 + 6 H2O

getting monosaccharides for glycolysis

glucose can be transported into the cell glucose molecules can be cleaved from glycogen or starch - yielding glucose 1-phosphate - converted to glucose 6-phosphate disaccharides can be hydrolyzed - maltose = two glucose - lactose = glucose and galactose - sucrose = glucose and fructose fructose, galactose, and mannose enter glycolysis at different points Entry of dietary glycogen, starch, disaccharides, and hexoses into the preparatory stage of glycolysis.

Central Importance of Glucose

glucose is an excellent fuel - yields good amount of energy upon oxidation - can be efficiently stores in the polymeric form - many organisms and tissues can meet their energy needs using only glucose glucose is a versatile biochemical precursor - bacteria can use glucose to build the carbon skeletons of : all the amino acids membrane lipids nucleotides in DNA and RNA cofactors needed for metabolism

Which product of Fructose-1phosphate aldolase requires triose kinase?

glyceraldehyde

continued synthesis of straight chain of glycogen

glycogen synthase continues synthesis of straight chains of glucose after 1st 8 glucose monomers are attached to glycogenin continues to use UDP-glucose to attach glucose to chain with an alpha1-4 a1-4 linkage until a chain of at least 11 monomers is made

making branches

glycogen-branching enzyme makes branches - transfers 6-7 glucose monomers from growing chain to c6 hydroxyl group of a glucose on the same chain or another chain - makes a1-6 alpha1-6 connection glycogen synthase can add more glucose monomers to both branches

Glycogen needs a starter molecule

glycogenin - enzyme - forms a protein core of glycogen molecule - catalyzes addition of first 8 glucose residues to itself using UDP-glucose

glycolysis overview

glycolysis was one of the earlies energy-yielding pathways it developed before photosynthesis when the atmosphere was still anaerobic thus, the task for early organisms was: how to extract free energy from glucose anaerobically the solution: - first: activate it by phosphorylation - second: collect energy from the high-energy metabolites glycolysis has 3 irreversible reactions - very negative deltaG' - point of regulation so glycolysis only goes forward when necessary - allosteric and reversible covalent regulation glycolysis occurs in the CYTOSOL of the cell

Enzymes = carry out reactions at physiological conditions so they proceed in a timely manner enzymes speed up the rate at which reaction proceeds towards its final equilibrium a typical EXERGONIC reaction : A + B AB*t C + D *t transition-state complex resembles both substrates and the products enzymes lower activation energy - energy required to form transition-state complex - they speed up reaction by lowering Ea How? by increasing concentrations of substrates at active site of enzyme by orienting substrates properly with respect to each other in order to form the transition-state complex , two models for enzyme-substrate interaction lock and key and induced fit

graph: frequency vs progress of reaction Ea { AB*t A + B ----------------- deltaG'o { C + D ------------------- reaction without enzyme = large peak at Ea reaction with enzyme = small peak at Ea

Name the enzymes that perform the 3 irreversible reactions in glycolysis.

hexokinase phosphofructokinase-1 pyruvate kinase

Fructose

hexokinase + ATP can make fructose 6-phosphate OR fructokinase + ATP can make fructose 1-phosphate fructose 1-phosphate aldolase cleaves fructose 1-phosphate - make dihydroxyacetone phosphate and glyceraldehyde ( NO phosphate ) triose kinase + ATP phosphorylates glyceraldehyde so it can enter glycolysis both routes require the same 2 ATP that glucose requires in preparatory phase

phosphoryl transfer from ATP

human mitochondria produce about 65 kg ATP/day to meet basic metabolic demands Phosphoryl group transfers: some of the participants. (c) When a nucleophile Z (in this case, the —OH on C-6 of glucose) attacks ATP, it displaces ADP (W) In this SN2 reaction, a pentacovalent intermediate (d) forms transiently.

hydrolysis of fats yields fatty acids and glycerol

hydrolysis of triacylglycerols is catalyzed by lipases triglyceride -- H2O / lipase --> diglyceride + fatty acid -- H2O / lipase --> monoglyceride + fatty acid

glucose transporter in membrane

hydrophobic outside polar inside glc inside 12 transmembrane helices Membrane topology of the glucose transporter GLUT1. (a) Transmembrane helices are represented here as oblique (angled) rows of three or four amino acid residues, each row depicting one turn of the α helix. Nine of the 12 helices contain three or more polar or charged residues (blue or red), often separated by several hydrophobic residues (yellow). This representation of topology is not intended to depict three-dimensional structure. (b) A helical wheel diagram shows the distribution of polar and nonpolar residues on the surface of a helical segment. The helix is diagrammed as though observed along its axis from the amino terminus. Adjacent residues in the linear sequence are connected, and each residue is placed around the wheel in the position it occupies in the helix; recall that 3.6 residues are required to make one complete turn of the α helix. In this example, the polar residues (blue) are on one side of the helix and the hydrophobic residues (yellow) on the other. This is, by definition, an amphipathic helix. (c) Sideby-side association of four amphipathic helices, each with its polar face oriented toward the central cavity, can produce a transmembrane channel lined with polar (and charged) residues. This channel provides many opportunities for hydrogen bonding with glucose as it moves through.

Membrane Dynamics: Lateral Diffusion

individual lipids undergo fast lateral diffusion within the leaflet very fast Motion of single phospholipids in a bilayer. (b) lateral diffusion within the leaflet is very rapid, requiring no catalysis.

What is the initial electron donor and final electron acceptor in Complex IV?

initial donor is cytochrome c and final electron acceptor is oxygen

Glycolysis vs. Gluconeogenesis

opposing pathways that are both thermodynamically favorable - operate in opposite directions > end product of one is the starting comopound of the other - both favorable > deltaG' = -16 kJ/mol overall for gluconeogenesis > deltaG' = -63 kJ/mol overall for glycolysis'' reversible reactions are used by BOTH pathways irreversible reaction of glycolysis must be bypassed in gluconeogenesis - highly thermodynamically favorable and regulated reactions - different enzymes in the different pathways - differentially regulated to prevent a futile cycle in eukaryotes, gluconeogenesis occurs in mitochondria and cytosol

structure of mitochondrion

outer membrane, inter membrane space, inner membrane, cristae (folds in im), matrix Biochemical anatomy of a mitochondrion. (a) The outer membrane has pores that make it permeable to small molecules and ions, but not to proteins. The convolutions (cristae) of the inner membrane provide a very large surface area. The inner membrane of a single liver mitochondrion may have more than 10,000 sets of electron-transfer systems (respiratory chains) and ATP synthase molecules, distributed over the membrane surface. (b) The mitochondria of heart muscle, which have more profuse cristae and thus a much larger area of inner membrane, contain more than three times as many sets of electron-transfer systems as (c) liver mitochondria. Muscle and liver mitochondria are about the size of a bacterium—1 to 2 μm long. The mitochondria of invertebrates, plants, and microbial eukaryotes are similar to those shown here, but with much variation in size, shape, and degree of convolution of the inner membrane.

Fructose has 2 potential pathways to enter glycolysis (one enters at fructose-6phosphate and the second enters at dihydroxyacetone phosphate/glyceraldehyde-3 phosphate). By the end of the prepatory phase, 2 ATP have been used as a part of both pathways. For each pathway, what are the 2 products of the phosphorylation reactions that use ATP?

pathway 1- fructose-6P and fructose-1,6bisP pathway 2- fructose-1P and glyceraldehyde-3P extra information ( not needed) : One pathway is a hexokinase can phosphorylate fructose to make fructose-6P. Fructose-6P is then phosphorylated by phosphofructokinase-1 to make fructose-1,6-biphosphate. The second pathways is fructose is phosphorylated by phosphofructokinase to make fructose-1P. Fructose 1P is then cleaved by fructose-1P-aldolase to make dihydroxyacetone phosphate and glyceraldehyde. That glyceraldehyde is phosphorylated by triose kinase to make glyceraldehyde-3P.

What other enzyme is required to allow this product to be used in glycolysis?

phosphoglucomutase which converts glucose 1-P to glucose 6-P.

Starting with pyruvate, which enzymes produce NADH? (hint, there are 4)

pyruvate dehydrogenase , isocitrate dehydrogenase , alpha-ketoglutamate dehydrogenase , malate dehydrogenase

Step 2 : phosphohexose isomerization

rationale - C1 of fructose is easier to phosphorylate in next step - allows for symmetrical cleavage by aldolase in Step 4 slightly thermodynamically unfavorable / reversible - product concentration kept low to drive forward

Step 5 : triose phosphate interconversion

rationale - allows glycolysis to proceed by one pathway aldolase creates two triose phosphates: - dihydroxyacetone phosphate ( DHAP) - glyceraldehyde 3-phosphate only glyceraldehyde-3P is the substrate for the next enzyme DHAP must be converted to glyceraldehyde-3P completes preparatory phase thermodynamically unfavorable / reversible - glyceraldehyde-3P kept low to pull reaction forward

Step 9: Dehydration of 2-PG to PEP dehydration of 2-phosphoglycerate to phosphoenolpyruvate

rationale - generate a high-energy phosphate compound hydrolysis of phosphate from 2-phosphoglycerate is NOT favorable enough - two negative charges in 2-PG are fairly close - but loss of phosphate from 2-PG would give a SECONDARY alcohol with NO further stabilization ( so change in free energy would not be very large) slightly thermodynamically unfavorable / reversible - product concentration kept low to pull forward

Step 6 : oxidation & phosphorylation of glyceraldehyde-3P

rationale - generation of a high-energy phosphate compound - incorporates inorganic phosphate - allows for production of ATP via glycolysis in next step oxidation of aldehyde ( loss of hydride ) with reduction of NAD+ and NADH phosphorylation makes a phosphorylated carboxyl group thermodynamically unfavorable / reversible - coupled to next reaction to pull forward

Step 7 : 1st Production of ATP

rationale - substrate level phosphorylation to make 2 ATP 1,3-bisphosphoglycerate is a high-energy compound - can donate the phosphate group to ADP to make ATP kinases are enzymes that transfer phosphate groups from ATP to various substrates - Phosphoglycerate kinase named for reverse reaction in gluconeogenesis highly thermodynamically favorable / reversible - is reversible because of couple to glyceraldehyde-3P dehydrogenase reaction

STEP 1 : phosphorylation of glucose

rationale - traps glucose inside the cell - no transporter for glucose 6-P - lowers intracellular glucose concentration to allow further transport by the glucose transporter this process uses the energy of ATP enzyme - hexokinase in eukaryotes - glucokinase in prokaryotes nucleophilic oxygen at C-6 of glucose attacks the last (y) phosphate of ATP highly thermodynamically favorable / irreversible

Step 8 : migration of the phosphate

rationale - be able to form high-energy phosphate compound mutases catalyze the change in position of functional groups within the molecule enzyme phosphorylated donates its phosphate to C-2 before removing the phosphate at C-3 - 2,3-bisphosphoglycerate intermediate - note that the phosphate from the substrate ends up bound to the enzyme at the end of the reaction thermodynamically unfavorable / reversible - reactant concentration kept high by phosphoglycerate kinase to push phosphoglycerate mutase reaction forward

STEP 4 : aldol cleavage of fructose 1,6-bisP

rationale - cleavage of six-carbon sugar into two three-carbon sugars - three-carbon sugars are high energy phosphate sugars thermodynamically unfavorable / reversible - glyceraldehude-3P concentration kept low to pull reaction forward - dihydroxyacetone phosphate converted to glyceraldehyde-3P in a separate reaction so product concentrations low

Step 10: 2nd Production of ATP

rationale - substrate-level phosphorylation to make ATP - net production of 2 ATP / glucose loss of phosphate from phosphoenolpyruvate yields an enol that tautomerizes into a ketone tautomerization - changes molecule with high free energy to molecule with low free energy > negative deltaG - effectively lowers the concentration of the reaction product - drives the reaction toward ATP formation highly thermodynamically favorable / irreversible - regulated by ATP, divalent metals, and other metabolites

animals and some bacteria undergo lactic acid fermentation

reduction of pyruvate to lactate - REVERSIBLE during strenuous exercise , lactate builds up in the muscle - oxygen doesnt reach muscle cells fast enough - generally less than 2 minutes of anaerobic exercise is possible the acidification of muscle prevents its continuous strenuous work the lactate can be transported to the liver and converted to glucose there -requires a recovery time - high amount of oxygen consumption to make ATP to allow gluconeogenesis to make glucose from lactate - glucose sent back to muscle > restores muscle glycogen stores erythrocytes ( NO mitochondria ) do lactic acid fermentation too

Ribulose and Xylulose are epimers

ribose is the aldehyde and ribulose is the ketone CH2OH-C=O-CHOH-CHOH-CH2OH D-Ribulose (H-C-OH for both) CH2OH-C=O-OHCH-CHOH-CH2OH D-Xylulose (OH-C-H and H-C-OH)

glycolysis importance

sequence of enzyme-catalyzed reactions by which glucose is converted into pyruvate - pyruvate can be further aerobically oxidized - pyruvate can be converted anaerobically to reduced molecules to recycle NADH to NAD+ - pyruvate can be used as a precursor in biosynthesis some of the free energy from oxidation is captured by the synthesis of ATP and NADH - NADH can help produce ATP through oxidative phosphorylation if oxygen is available

citric acid cycle overview

stage 1 acetyl-CoA production generates ATP, NADH, FADH2 depending on the pathway stage 2 acetyl-CoA oxidation generates GTP and more NADH and FADH2 stage 3 electron transfer and oxidative phosphorylation generates a lot of ATP

b-oxidation pathway steps

step 1: dehydrogenation of alkane to alkene Catalyzed by isoforms of acyl-CoA dehydrogenase (AD) - Long-chain AD (12-18 carbons)- in inner membrane - Medium-chain AD (4-14 carbons)- in matrix - Short-chain AD (4-8 carbons)- in matrix Transfer of 2 H+ + 2 e- from fatty acyl-CoA to FAD results in trans double bond, different from naturally occurring unsaturated fatty acids Electrons from bound FADH2 transferred to the electron transport chain by the mobile carrier electron-transferring flavoprotein (ETF) ETF binds to ETF:Q oxidoreductase which also contains FAD and ultimately reduces ubiquinone Step 2: hydration of alkene Catalyzed by two isoforms of enoyl-CoA hydratase: - Soluble short-chain hydratase for fatty acids less than 12 C long - Membrane-bound long-chain hydratase, part of Trifunctional protein Water added across the double bond yielding alcohol

four major pathways of glucose utilization

storage - can be stored in the polymeric form ( starch and glycogen) - when glucose and ATP high glycolysis - generates energy via oxidation of glucose > make ATP - short-term energy needs - can link to other pathways to generate more energy pentose phosphate pathway - generates NADPH via oxidation of glucose - for detoxification and the biosynthesis of lipids and nucleotides synthesis of structural polysaccharides - for example, in cell walls of bacteria, fungi, and plants

synthesis of branches in glycogen

synthesis in glycogen The glycogen-branching enzyme also called amylo (1-4) to (1-6) transglycosylase , or glycosyl-(4-6) transferase forms a new branch point during glycogen synthesis

synthesis of oxaloacetate

synthesis of phosphoenolpyruvate from pyruvate. (a) In mitochondria, pyruvate is converted to oxaloacetate in a biotin-requiring reaction catalyzed by pyruvate carboxylase

Why does the cell keep ATP high?

the actual ΔG for the hydrolysis of the phosphate group is even more favorable (or more negative) than the standard ΔG. You need to indicate the reaction for which the actual ΔG changes. Also, if the actual ΔG is higher, then it would be less favorable. It is more negative when [ATP] is kept high. (it's hard to say it right) (The ratio of [ADP][Pi]/ [ATP] will be low in the second term of the ΔG equation making the whole term negative)

anaerobic glycolysis

the overall process where pyruvic acid generated during glycolysis is converted to lactic acid

In the payoff phase of glycolysis, what it the payoff?

the payoff phase produces 2 NADH, 2 pyruvate, and 4 ATP.

What are the products of each of the 2 steps of the first bypass and which of them must be produced in mitochondria?

the products of the first step of the first bypass reaction are oxaoloacetate, ADP, and Pi. This step must occur in the mitochondria because the enzyme used to catalyze this reaction, pyruvate carboxylase, is located there The products of the second step are phosphoenolpyruvate, GDP, and CO2 This reaction can occur in either the cytosol or mitochondria, depending on the organism.

iron-sulfur clusters

this prosthetic group is present in complex I, II, and III of electron transport - One electron carriers-Fe3+ to Fe2+ - Coordinated by cysteines in the protein-stationary - Contain variable numbers of iron and sulfur atoms Iron-sulfur centers. The Fe-S centers of iron-sulfur proteins may be as simple as (a), with a single Fe ion surrounded by the S atoms of four Cys residues. Other centers include both inorganic and Cys S atoms, as in (b) 2Fe-2S or (c) 4Fe-4S centers. (d) The ferredoxin of the cyanobacterium Anabaena 7120 has one 2Fe-2S center (PDB ID 1FRD); Fe is red, inorganic S is yellow, and the S of Cys is orange. (Note that in these designations only the inorganic S atoms are counted. For example, in the 2Fe-2S center (b), each Fe ion is actually surrounded by four S atoms.) The exact standard reduction potential of the iron in these centers depends on the type of center and its interaction with the associated protein.

Yeast undergo ethanol fermentation

two step reduction of pyruvate to ethanol production of acetaldehyde from pyruvate is IRREVERSIBLE humans do not have pyruvate decarboxylase we do express alcohol dehydrogenase for ethanol metabolism ( runs in reverse of fermentation) CO2 produced in the first step is responsible for - carbonation in beer - dough rising when baking bread

3 classes of transport systems

uniport, symport, antiport cotransport = symport and antiport Cotransport involves 2 or more molecules Three general classes of transport systems. Transporters differ in the number of solutes (substrates) transported and the direction in which each solute moves. Examples of all three types of transporter are discussed in the text. Note that this classification tells us nothing about whether these are energy-requiring (active transport) or energy-independent (passive transport) processes.

Glycogen synthesis

when glucose 6-phosphate is high in the cell then it is helpful to control osmotic pressure by synthesizing glycogen ** but only when ATP is high glucose 6-P is converted to glucose 1-P by the enzyme Phosphoglucomutase glucose 1-P is then activated by recreating with UTP to make UDP-glucose in a reaction catalyzed by UDP-glucose Pyrophosphorylase pyrophosphate is the leaving group UMP is attached to phosphate of glucose 1-P reaction made favorable by the breakdown of pyrophosphate to 2Pi ( PPi ) by inorganic pyrophosphatase

thermodynamics of membrane transport

ΔG of Transport for Uncharged Molecules deltaG ΔGt= RTln (C2/C1) C1 is the concentration of the molecule on the side of the membrane where it starts C2 is the concentration of the molecule on the side of the membrane where the molecule will be moved to If the molecule goes from a region of high concentration to low concentration, then the ΔGt will be negative. Transport is favorable If the molecule goes from a region of low concentration to a region of high concentration, then it will not be favorable and transport will require and input of energy to occur. ΔG of Transport for Charged Molecules deltaG ΔGt= RTln (C2/C1) + zFΔψ Transport of charged molecules depends on both the concentration on each side of the membrane and the charge of the molecule as well as the charge on the membrane (one side of the membrane will be positive and one will be negative) C2 and C1 are the same as for uncharged molecules z is the charge on the molecule including both the sign and the number. Mg2+ would have a z of +2. Cl- would have a z of -1. F-Faraday constant 96.5 kJ/molV Δψ is the charge on the membrane. It also has a sign. If the molecule is moving to the positive side of the membrane, then the sign will be + . If the molecule is moving to the negative side of the membrane, then the sign will be -. If a positively charged molecule (z is +), is moving to the negative side of the membrane (Δψ is -), then the second term will be negative. So the charge portion of the transport will be favorable. This makes sense because the molecule is moving down the charge gradient. If a positively charged molecule (z is +), is moving to the positive side of the membrane (Δψ is +), then the second term will be positive. So the charge portion of the transport will be unfavorable. This makes sense because the molecule is moving up the charge gradient. However, in either case, the concentration term also contributes to the determination of whether transport will be favorable

electron transport is coupled to ATP synthesis

• As described, ATP synthesis requires electron transport • But electron transport also requires ATP synthesis CN blocks e- transfer to O2 oligomycin inhibits ATP synthesis dinitrophenol uncouples reactions- equalizes H+ concentration across membrane Coupling of electron transfer and ATP synthesis in mitochondria. In experiments to demonstrate coupling, mitochondria are suspended in a buffered medium and an O2 electrode monitors O2 consumption. At intervals, samples are removed and assayed for the presence of ATP. (a) Addition of ADP and Pi alone results in little or no increase in either respiration (O2 consumption; black) or ATP synthesis (red). When succinate is added, respiration begins immediately and ATP is synthesized. Addition of cyanide (CN-), which blocks electron transfer between cytochrome oxidase (Complex IV) and O2, inhibits both respiration and ATP synthesis. (b) Mitochondria provided with succinate respire and synthesize ATP only when ADP and Pi are added. Subsequent addition of venturicidin or oligomycin, inhibitors of ATP synthase, blocks both ATP synthesis and respiration. Dinitrophenol (DNP) is an uncoupler, allowing respiration to continue without ATP synthesis.

Chemiosmotic Model for ATP synthesis

• Electron transport sets up a proton-motive force • Energy of proton-motive force drives synthesis of ATP Chemiosmotic model. In this simple representation of the chemiosmotic theory applied to mitochondria, electrons from NADH and other oxidizable substrates pass through a chain of carriers arranged asymmetrically in the inner membrane. Electron flow is accompanied by proton transfer across the membrane, producing both a chemical gradient (∆pH) and an electrical gradient (∆ψ) (combined, the proton-motive force). The inner mitochondrial membrane is impermeable to protons; protons can reenter the matrix only through proton-specific channels (Fo). The protonmotive force that drives protons back into the matrix provides the energy for ATP synthesis, catalyzed by the F1 complex associated with Fo.