Cell bio chap. 10 lecture

For each electron pair delivered to the ETS by NADH, 10H+ are pumped into the inter membrane space; if 3 are required to make one ATP molecule, approximately how many ATPs are made per NADH and FADH2?

-3 for NADH -2 for FADH2 (protons pumped into the inter membrane space: 6)

How many protons are pumped from the matrix to the inter membrane space at each complex of the electron transport chain?

-4 H+ are pumped out of the matrix by complex 1 -Succinate-coenzyme Q (complex 2) carries 2 H+ across the inner membrane and 2 more H+ are pumped out of the matrix (total of 4 from complex 3) -2 more H+ are pumped from the matrix by complex 4

How is the electrochemical gradient harnessed to produce ATP? (what is the structure of the ATP synthase enzyme?)

-ATP synthesis: the F0F1 ATP synthase lollipop: a miniature motor in which proton flow turns a microscopic gear that drives ATP synthesis -F0= channel for proton flow -F1= carries out synthesis of the ATP

Does NADH inhibit or stimulate the citric acid cycle? ADP?

-Allosteric enzymes -Increased NADH inhibits the citric acid cycle (specifically pyruvate dehydrogenase) -ADP stimulates the citric acid cycle (ATP is formed from ADP)

What is an electrochemical gradient?

-An electrochemical gradient is a gradient of electrochemical potential, usually for an ion that can move across a membrane. The gradient consists of two parts, the chemical gradient, or difference in solute concentration across a membrane, and the electrical gradient, or difference in charge across a membrane -Proton gradients are high in the inter membrane space of mitochondria and low in the matrix -Eventually, the hydrogen ions (protons) from the inter membrane space are passed to be pumped from the ATP synthase enzyme to synthesize ATP (3 H+ come from inter membrane space to do this)

Coenzyme Q (ubiquinone)

-One of the 5 electron carriers of the ETS -ONLY LIPID -Accepts hydrogen atoms -Not a protein

Iron sulfur proteins

-One of the five carriers of the ETS -Proteins-iron-sulfur in middle -Accept electrons -Fe3+ + electron --> Fe2+

Copper containing cytochromes

-One of the five electron carriers of the ETS -Protein -Cu2+ (cupric) + electron --> Cu1+ (cuprous)

Number of oxidations, NADH produced, number of FADH2 produced, ATP produced, and decarboxylation reactions in one "lap" of the citric acid cycle

-Oxidation reactions: 4 -NADH produced: 3 -FADH2 produced: 1 -ATP produced: 1 -Decarboxylation reactions: 2

Number of oxidations, NADH produced, number of FADH2 produced, ATP produced, and decarboxylation reactions in two "laps" of the citric acid cycle

-Oxidation reactions: 8 -NADH produced: 6 -FADH2 produced: 2 -ATP produced: 2 -Decarboxylation reactions: 4

What are the alternate names for the Krebs cycle?

-Tricarboxylic acid -Citric acid cycle

Cytochromes

-Vinyl groups -One of the five electron carriers of the ETS -Protein -Heme group -Fe3+ (ferric) + electron --> Fe2+ (ferrous) -Porphoryn ring

How many ATP molecules are formed per electron pair from FADH2?

2

How many ATP are estimated to be generated for each electron pair of FADH2?

2 ATP estimated to be generated

How many decarboxylation reactions occur during the Krebs cycle?

2 for one lap, 4 for 2 laps

Why is the Krebs cycle referred to as a CYCLE?

2 laps of the cycle: one for each acetyl CoA

How many "laps" of the Krebs cycle are necessary when you start with a single glucose molecule?

2 laps, one for each acetyl CoA produced (2 acetyl CoA produced after pyruvate oxidation, glucose was originally split in half at the beginning of glycolysis)

How many laps are there of the citric acid cycle?

2 laps; one lap for each acetyl-CoA

What events the place in the mitochondria?

2. Pyruvate oxidation: pyruvate --> acetyl CoA; then continues to Citric Acid Cycle 3. Citric acid cycle (Krebs cycle) 4. Electron transport chain and proton pumping 5. ATP synthesis

How many ATP molecules are formed per electron pair from NADH?

3

How many ATP are estimated to be generated for each electron pair from NADH?

3 ATP estimated to be generated

How many NADH are produced from the Krebs cycle?

3 for one lap, 6 for 2 laps

How many protons are needed to produce 1 ATP by ATP synthase?

3 protons (H+) move through "a" channel for each ATP made

Products of the Krebs cycle

4 CO2, 6 NADH, 2 FADH2, & 2 ATP -NADH & FADH2 can be used to form additional ATP through the electron transport chain

How many oxidation reactions occur in the Krebs cycle?

4 for one lap, 8 for 2 laps

What is F0F1 ATPase?

Enzymes that make ATP

Cristae

Folds of inner membrane of mitochondria

Where do the electrons that are delivered to the electron transport chain come from?

From NADH and FADH2

Positive reduction potentials

Good electron acceptors

Negative reduction potentials

Good electron donors

Outer membrane properties of mitochondria

Highly permeable; contains porins (channels); and allows any solute < 10,000 MW IN

What is the ENDOSYMBIONT THEORY and the evidence that supports it?

Mitochondrion was a bacterium engulfed by a eukaryotic cell

How are most of the electron carriers arranged?

Most of the electron carriers are arranged in 4 large complexes on the cristae

Substrate level phosphorylation

Substrate-level phosphorylation is a metabolic reaction that results in the formation of ATP or GTP by the direct transfer of a phosphoryl (PO3) group to ADP or GDP from another phosphorylated compound

Where in the cell does pyruvate oxidation take place?

Takes place in the mitochondrial matrix

Where in the cell does the citric acid cycle occur?

Takes place in the mitochondrial matrix

How is the efficiency of glucose oxidation determined? (equation)

(ATP generated per glucose molecule (38) x delta G for ATP hydrolysis (-10kcal mol)) / delta G for complete catabolism of glucose (-686 kcal/mol) Multiplied by 100

Explain how F0 and F1 of ATP synthase result in ATP formation (and explain the structure)

-"a" is located on the side of the ring of "c"s; a "gamma" subunit is located under the ring of cs which is attached to an alpha-beta ring -F0 is located in the cristae below the inter membrane space -F1 is located in the matrix -Protons flow through the "a" channel in the F0 subunit ("downhill") (3 H+ move through "a" channel for each ATP made) -This causes the ring of 10 c subunits in the F0 portion to rotate -As a result, the gamma subunit of the F1 subunit spins -This activates the alpha-beta catalytic ring of the F1 subunit where ADP and P bind to form ATP

In which direction are protons pumped in the chemiosmotic coupling model?

-10 protons (H+) are pumped from the matrix to the intermembrane space via complexes 1, 3, and 4 -3 protons (H+) are pumped from the inter membrane space to the matrix via ATP synthase to activate it to make one ATP from ADP + P

Pyruvate oxidation (what is reduced and oxidized?)

-2 pyruvates undergo decarboxylation (CO2 produced) -Pyruvate oxidized to acetyl CoA

Second step of citric acid cycle

-An enzyme called aconitase converts citrate into isocitrate. Next, an isocitrate dehydrogenase enzyme oxidizes isocitrate, a six-carbon molecule, to a five-carbon α-ketoglutarate (the removal of CO2 rearranges the structure). The carbon that was lost is released as carbon dioxide and one NADH is also formed (NADH + H+). The carbon dioxide that is released was originally part of oxaloacetate and not acetyl CoA **OXIDATION AND DECARBOXYLATION**

The five electron carriers of the ETS

-Can be oxidized or reduced (has prosthetic group, part of molecule oxidized or reduced) -Flavoproteins -Cytochromes -Iron sulfur proteins -Coenzyme Q (ubiquinone) -Copper containing cytochromes

Electron transport description

-Electron flow from reduced NAD and FAD to oxygen (final electron acceptor) -Electrons are passed from NADH and FADH2 generated in glycolysis, pyruvate oxidation, and Krebs cycle to a series of electron carriers called the electron transport system (ETC: electron transport chain)

Describe the organization of the four complexes of the electron transport chain. How do electrons flow between them?

-Electron transfer must occur downhill -Electrons can only be transferred to a molecule with a more positive reduction potential -Complex 1: NADH dehydrogenase -Complex 2: succinate-coenzyme Q oxidoreductase -Complex 3: coenzyme Q-cytochrome c oxidoreductase -Complex 4: cytochrome c oxidase

Do the 5 classes of carriers on the electron transport chain receive electrons or hydrogen atoms?

-FMN on flavoproteins receive hydrogen atoms -Heme (Fe) of cytochromes receive electrons (Fe3+ ferric --> Fe2+ ferrous) -Iron sulfur proteins accept electrons (Fe3+ --> Fe2+) -Copper containing cytochromes accept electrons (Cu2+ cupric --> Cu1+ cuprous) -Coenzyme Q (ubiquinone) accepts hydrogen atoms

Where is ATP synthase locate in the mitochondria?

-Fo complex (stalk) is embedded in the inner membrane -F1 complex is located in the matrix

What are the "lollipops" embedded in the inner mitochondrial membrane?

-FoF1 ATP synthase -Fo= stalk embedded in cristae -F1= complex in MATRIX -Enzymes that make ATP

How is water formed from the electron transport chain?

-Form H2O (metabolic H2O) -Formed by adding the 2 hydrogen ions to the final electron acceptor (1/2O2)

Net inputs and products of Krebs cycle

-Input: acetyl CoA -Products: NADH, FADH2, ATP

What is being oxidized and reduced in the Krebs cycle? Where do the electrons go?

-Isocitrate is oxidized (reducing NAD+ to NADH) -Alpha-ketogluterate is oxidized (reducing NAD+ to NADH) -Succinate is oxidized (reducing FAD to FADH2) -Malate is oxidized to form oxalacetate (reducing NAD+ to NADH) -Electrons go to oxidized forms of NAD and FAD

Outline of the electron carriers in the complexes (also linking molecules and how electrons from NADH and FADH2 are delivered to the complexes)

-NADH enters complex 1 with 2 electrons (NADH dehydrogenase); complex 1 comprises of many FMN (flavoprotein) --> FeS (iron sulfur protein) ; gives off ATP; gives off electron to linker ubiquinone -Ubiquinone is a linker molecule that is not in a complex (LIPID) and diffuses rapidly in a cristae -Complex 2 (succinate dehydrogenase-step in Krebs cycle) goes to linker ubiquinone with electrons; complex 2 comprises of FAD--> FeS; FADH2 released -Electrons from ubiquinone "pool" given to complex 3 (cytochrome b-c1 complex) ; comprises of Cyt b --> FeS --> Cyt C1 --> linker cyt-c; releases ATP -Cyt c is a linker molecule between complexes 3 and 4 -Electrons from linker molecule Cyt-c enter complex 4 (cytochrome c oxidase); comprises of cyt a --> cyt a3 --> O2; ATP is released -O2 is the final electron accepter and eventually forms into H2O

Description of the structure of the mitochondria

-OUTER MEMBRANE: highly permeable; contains porins (channels); and allows any solute < 10,000 MW IN -INTERMEMBRANE SPACE: composition similar to cytosol -INNER MEMBRANE (FORTRESS): impermeable; transport proteins present; permeability barrier; contains phospholipids UNIQUE TO BACTERIA -MATRIX: contains bacterial DNA and bacterial ribosomes -CRISTAE: folds of inner membrane

Description of the chemiosmotic coupling model of the electron transport chain (outline of events)

-Peter Mitchell (1961) -Key feature: proton translocation (move hydrogen ions/protons across the cristae) -Establishment of an ELECTROCHEMICAL PROTON (H+) GRADIENT (proton-motive force) 1) An electron pair is delivered to the ETS by NADH and passed via complexes 1, 3, and 4 to oxygen, the final electron acceptor to produce water 2) Hydrogen ions (protons) are pumped through complexes 1, 3, and 4 from the matrix to the intermembrane space Complex 1: pumps 4H+ Ubiquinone and complex 3: pump 4H+ Complex 4: pumps 2H+ TOTAL of 10 protons 3) Hydrogen ions (protons) accumulate in the intermembrane space, lowering the pH, and creating an electrochemical proton gradient (making the inter membrane space acidic) 4) 3 hydrogen ions pass down their electrochemical gradient through the enzyme ATP synthase, activating it, to make one ATP from ADP and P

Flavoproteins (prosthetic group and general structure)

-Prosthetic group: Flavin mononucleotide (FMN) -Hydrogen ions connect to "N" on nucleotides -One of the five electron carriers of the ETS

Are the five classes of carriers on the electron transport chain proteins or lipids?

-Proteins: flavoproteins, cytochromes, copper containing cytochromes, and iron sulfur proteins -ONLY LIPID: coenzyme Q (ubiquinone)

What is reduction potential?

-Referring to the electron transport chain -A measure in VOLTS of the affinity of a compound for electrons -Reduction potential=(E0')

Electron shuttle systems

-Solution to the problem of eukaryote's inability for NADH to penetrate the mitochondria inner membrane -In the outer mitochondria membrane (cytosol), NADH from glycolysis is oxidized to NAD+ to heart, kidney, liver -2 NADH in the cytosol of the outer mitochondria membrane are oxidized to NAD+ to the muscle, brain and other tissues -With a shuttle system located at the inner mitochondria membrane, electrons donated from the outer mit. membrane form 2NADH at the inner mitochondria membrane which in turn make 6 ATP; the electrons from the oxidized NAD+ that go to the muscle, brain, and other tissues are also donated to the shuttle system located at the inner membrane of the mitochondria to make FADH2 which in turn form 4 ATP

First step of the citric acid cycle

-The cycle begins with the entry of the acetate group of acetyl CoA produced in pyruvate oxidation -In the first step of the cycle, an enzyme called CITRIC SYNTHASE joins the 2-Carbon acetyl group from the acetyl CoA with a 4-Carbon oxaloacetate to form a 6-Carbon citrate (tricarboxylic acid)

Why does the maximum theoretical ATP yield vary between 36 and 38 in eukaryotic cells?

-The problem: with eukaryotes, NADH generated in glycolysis cannot penetrate the mitochondria inner membrane -Therefore, the solution is electron shuttle systems -In prokaryotic cells, ATP generated is always 38

Why are the electron transport chain carriers in a particular sequence?

-There is a decrease in free energy as electrons pass along the chain to oxygen -The electron carriers function in a sequence determined by their reduction potential

How many FADH2 are produced from the Krebs cycle?

1 for one lap, 2 for 2 laps

How many ATP are formed from the Krebs cycle and by what process?

1 for one lap, 2 for 2 laps -Substrate level phosphorylation

At which complexes is ATP formed of the electron transport chain?

1, 3, and 4 (no ATP formed at complex 2)

Glucose catabolism: adding up the ATP generated (outline)

ATP FORMED GLYCOLYSIS: 2 PYRUVATE OXIDATION: 0 KREBS CYCLE: 2 NADH GENERATED GLYCOLYSIS: 2 PYRUVATE OXIDIATION: 2 KREBS CYCLE: 6 ATP GENERATED PER NADH: 3 THEREFORE, TOTAL ATP FROM NADH: 30 FADH2 GENERATED GLYCOLYSIS: 0 PYRUVATE OXIDATION: 0 KREBS CYCLE: 2 ATP GENERATED PER FADH2: 2 THEREFORE, TOTAL ATP FROM FADH2: 4 MAXMUM THEORETICAL TOTAL ATP GENERATED FROM THE CATABOLISM OF ONE GLUCOSE MOLECULE: 38

Oxidative phosphorylation

ATP synthesis using the energy released during electron transport

How much of the energy in a glucose molecule is captured to synthesize ATP? How efficient is it?

About 50%, highly efficient

Are the electron carries of the electron transport chain left in a reduced or oxidized form?

Carriers are left oxidized

Where does oxidative phosphorylation occur in a bacterium?

Cell membrane

Porins

Channels located on outer membrane of mitochondria

Inner membrane space properties of mitochondria

Composition similar to cytosol

Inner membrane properties of mitochondria

Impermeable; transport proteins present; permeability barrier; contains phospholipids UNIQUE TO BACTERIA

Step 5 of the citric acid cycle

In step five, a succinyl CoA synthetase enzyme converts succinyl CoA (4C) into succinate (4C). This produces GTP (P was added to GDP) which is converted to ATP. (in an animal)

Step 7 of the citric acid cycle

In step seven, a fumarate hydratase enzyme then converts fumarate into malate (4C)

Step 6 of the citric acid cycle

In step six, an enzyme called succinate dehydrogenase converts succinate (4C) into fumarate by giving its electron away (oxidation). This step makes one FADH2. **SUBSTRATE LEVEL PHOSPHORYLATION AND ATP PRODUCTION**

Step 8 of the citric acid cycle (final step)

In the final step of the citric acid cycle, a malate dehydrogenase enzyme converts malate (4C) back to oxaloacetate (4C) by oxidation. Like all steps involving a dehydrogenase, a coenzyme is produced. Here it is NADH (NAD+ was reduced to NADH)

The citric acid cycle (ALL STEPS) shit is long

In the first step of the cycle, an enzyme called citrate synthase joins the two-carbon acetyl group from acetyl CoA with the four-carbon oxaloacetate to form a six-carbon citrate. In step two, an enzyme called aconitase converts citrate into isocitrate. Next, an isocitrate dehydrogenase enzyme oxidizes isocitrate, a six-carbon molecule, to a five-carbon α-ketoglutarate. The carbon that was lost is released as carbon dioxide and one NADH is also formed. The carbon dioxide that is released was originally part of oxaloacetate and not acetyl CoA. In the fouth step, an enzyme called α-ketoglutarate dehydrogenase converts α-ketoglutarate into a four-carbon succinyl CoA. Similar to step three, this reaction produces one carbon dioxide and one NADH. In step five, a succinyl CoA synthetase enzyme converts succinyl CoA into succinate. This produces GTP which is converted to ATP. In step six, an enzyme called succinate dehydrogenase converts succinate into fumarate. This step makes one FADH2. In step seven, a fumarate hydratase enzyme then converts fumarate into malate. In the final step of the citric acid cycle, a malate dehydrogenase enzyme converts malate back to oxaloacetate. Like all steps involving a dehydrogenase, a coenzyme is produced. Here it is NADH. The oxaloacetate that was regenerated through the citric acid cycle is now ready to join with another acetyl group and begin the cycle a second time. For every one glucose that is broken down through glycolysis, two pyruvates will be produced. These two pyruvates will produce two acetyl CoAs. So, for every one glucose, two acetyl CoAs will be made and two turns of the citric acid cycle will occur. This means each product of the cycle must be doubled.

The third/fourth step of citric acid cycle

In the fourth step, an enzyme called α-ketoglutarate dehydrogenase converts α-ketoglutarate into a four-carbon succinyl CoA. Similar to step three, this reaction produces one carbon dioxide and one NADH. -Oxidized NAD+ reduced to NADH + H+ **OXIDATION AND CARBOXYLATION**

Where is the electron transport chain located?

Inner mitochondrial membrane

Outline how the electron transport chain operates

NADH loses 2 electrons → NAD+ → oxidized FMN reduced to FMN2 → oxidized quinone carried by ubiquinone is reduced to hydroquinone → oxdized 2Fe3+ is carried by cytochrome b and reduced to 2 Fe2+ → reduced 2Fe2+ is carried by cytochrome c1 and oxidized to 2Fe3+ → oxidized 2Fe3+ is carried by cytochrome c and is reduced to 2Fe2+ → oxdized 2Fe3+ is carried by cytochrome a-a3 complex and reduced to 2Fe2+ → oxidized 2Cu2+ is carried by cytochrome a-a3 complex and reduced to 2Cu+ → 1/2O2 (final electron acceptor) + 2H+ to H2O (product)

Why are the numbers of ATP formed from electron pair of NADH and FADH2 different?

NADH produces 3 ATP during the ETC (Electron Transport Chain) with oxidative phosphorylation because NADH gives up its electron to Complex I, which is at a higher energy level than the other Complexes. When Complex I transfers the electron to Complex III, energy is given off to pump protons across the membrane, creating a gradient. The electron moves again to Complex IV and again pumps more electrons across the membrane. Because NADH started with Complex I, it had more chances to pumps more protons across the gradient, which powers the ATP synthase and gives us 3 ATP per molecule of NADH. FADH2 produces 2 ATP during the ETC because it gives up its electron to Complex II, bypassing Complex I. By bypassing Complex I, we missed a chance to pump protons across the membrane, so less protons have been pumped by the time we get to Complex IV. Protons still have been pumped, enough to fuel 2 ATP created by ATP synthase.

What is the 4C starting molecule that receives the acetyl group from acetyl CoA? (Krebs cycle)

Oxalacetate

How does reduction potential explain the order of the carriers in the electron transport chain?

The electron carriers function in a sequence determined by their reduction potential

The electron transport system (description)

The electron carriers of the chain receive electrons from NADH and FADH2; each carrier is successfully reduced by the electrons from the proceeding carrier in the chain and is oxidized by the loss of electrons to the carrier following it; thus electrons are passed from one carrier to the next like a "bucket brigade" until the final acceptor is reached

Aerobic respiration

The exergonic process by which cells oxidize glucose to carbon dioxide and water using oxygen as the final electron acceptor with a significant portion of the released energy conserved as ATP (Becker) C6H12O6 + 6O2 (final electron acceptor) --> 6CO2 + 6H2O + energy (captured to make ATP)

The citric acid cycle and catabolism of fats and proteins

The fatty acids are linked to coenzyme A to form fatty acyl CoAs, which are then degraded by beta-oxidation, a catabolic process that generates acetyl CoA and the reduced coenzymes NADH and FADH2

What is the first 6C product formed during the Krebs cycle?

Tricarboxylic acid (oxalacetate + citrate)

Summary of detailed events in establishing the electrochemical gradient

a) Complex 1: receives 2 electrons from NADH and passes them to CoQ via FMN (flavoproteins) and an FeS (iron sulfur) protein; during this process, 4H+ are pumped out of the matrix by complex 1 b) Complex 3 passes electrons from CoQH2 to cytochrome c via cytochromes b and c1 and an Fe-S protein (iron sulfur); CoHQ2 carries 2H+ across the inner membrane and 2 more H+ are pumped out of the matrix c) Complex 4 receives electrons from cytochrome c and, via cytochrome a and a3, passes them to molecular oxygen, which is reduced to water as 2 more H+ are pumped from the matrix by complex 4 d) ATP synthase uses the energy from the proton gradient generated during electron transport to synthesize ATP from ADP and P

Overall equation of the citric acid cycle

acetyl CoA + (3NAD+) + FAD + ADP + P ---> 2CO2 + 3NADH + FADH2 + CoA-SH + ATP