3rd Unit / Ch 9 TCA Krebs Cycle Flashcards
Pyruvate Dehydrogenase Complex 9.1
List the five coenzymes required by the nonregulatory enzymes of the PDHC. Do they function as prosthetic groups or as cosubstrates?
Of the five coenzymes required by the nonregulatory enzymes of the PDHC that catalyzes the oxidative decarboxylation of pyruvate, TPP of E1 ( pyruvate decarboxylase ), lipoic acid of E2 ( dihydrolipoyl transacetylase ), and FAD of E3 ( dihydrolipoyl
dehydrogenase ) are tightly bound and function as coenzyme- prosthetic groups. CoA and NAD+ (loosely associated with E2 and E3, respectively) are coenzymecosubstrates.
[Note: Arsenite inhibits lipoic acid– requiring enzymes.]
Pyruvate Dehydrogenase Complex 9.1
How do ATP, acetyl CoA, NADH, and pyruvate regulate the PDHC? How does Ca2+ regulate it?
ATP, acetyl CoA, and NADH ( PDHC products) are allosteric activators of PDH kinase. The kinase phosphorylates and inhibits E1 of the PDHC. Pyruvate ( PDHC substrate) is a potent inhibitor of the kinase. Ca2+, released during skeletal muscle contraction, is an allosteric activator of PDH phosphatase. The phosphatase
dephosphorylates and activates the PDHC, increasing ATP availability to power contraction. NADH and acetyl CoA also affect PDHC nonregulatory enzymes by product inhibition.
Pyruvate Dehydrogenase Complex 9.1
Why does a deficiency in PDHC activity cause lactic acidosis?
A deficiency of the PDHC results in a rise in pyruvate, thereby upregulating other reactions of pyruvate metabolism such as reduction to lactate by NADH-requiring
LDH leading to lactic acidosis.
[ Note: The most common form of PDHC
deficiency is caused by mutations to E1.]
Tricarboxylic Acid Cycle 9.2
What intermediate of the TCA (Krebs) cycle is combined with acetyl CoA to initiate the cycle (and is regenerated by the cycle), as shown?
Name the enzyme that catalyzes step 4 and compare it to the PDHC.
OAA is combined with acetyl CoA by Citrate Synthase to initiate the TCA cycle and is regenerated by the cycle.
The step 4 enzyme is the a -KGD complex , which catalyzes an oxidative decarboxylation reaction in a manner analogous to that of the PDHC. It is a protein aggregate of three nonregulatory enzymes that use the same coenzymes as the PDHC (their E3 components are identical). Like PDHC, its E2 is inhibited by
arsenic. Unlike PDHC, it is not regulated covalently.
Tricarboxylic Acid Cycle 9.2
Which step generates GTP? How?
Substrate-level phosphorylation of GDP to GTP occurs during the conversion of succinyl CoA to succinate (step 5) by succinate thiokinase via cleavage of the high-energy thioester bond in succinyl CoA.
Tricarboxylic Acid Cycle 9.2
What is unique about the enzyme that catalyzes step 6?
FAD-containing Succinate Dehydrogenase of step 6 is the only TCA cycle enzyme embedded in the inner mitochondrial membrane (the others are in the matrix). SD is a component of ETC Complex II.
Tricarboxylic Acid Cycle 9.2
Which TCA cycle enzyme will be directly affected with
thiamine ( vitamin B1 ) deficiency?
What encephalopathy–psychosis syndrome can develop with this deficiency?
The a - KGD complex of the TCA cycle will be directly inhibited by a thiamine ( vitamin B1 ) deficiency because of its requirement for TPP, a thiamine derivative. Wernicke-Korsakoff syndrome can develop with thiamine deficiency (e.g., in alcohol abusers). [Note: PDHC activity would also be inhibited with thiamine deficiency.]
Tricarboxylic Acid Cycle 9.3
How is the TCA cycle enzyme CS shown regulated?
CS, which catalyzes the first of three irreversible reactions of the TCA cycle, is nonallosterically inhibited by its
product citrate (a tricarboxylic acid).
Tricarboxylic Acid Cycle 9.3
What effect would elevated ATP and/or NADH levels have on Isocitrate dehydrogenase activity? Would Ca2+ have the same effect?
ATP and NADH (high-energy signals) are allosteric inhibitors of ICD, the second of three irreversible reactions of the cycle. In contrast, Ca2+ (and ADP) allosterically activates the enzyme. Both Ca2+ and ADP signal the need for energy.
Tricarboxylic Acid Cycle 9.3
Fluoroacetate is a plant toxin that is used as a pesticide. It inhibits aconitase of the TCA cycle. What effect will fluoroacetate poisoning have on aerobic metabolism.
Aerobic metabolism will be inhibited with fluoroacetate poisoning. By inhibiting aconitase, fluoroacetate will prevent NADH and FADH 2 production in the TCA cycle, thereby decreasing ATP production
by OXPHOS (and of GTP by substrate-level phosphorylation). Anaerobic metabolism will increase, providing
some ATP and causing a lactic acidosis.
Tricarboxylic Acid Cycle 9.4
What coenzyme is required for the Malate Dehydrogenase reaction shown?
What role does this reaction play outside of the cycle (e.g., in glycolysis)?
The required coenzyme is NAD as NAD+ in the forward reaction and NADH in the reverse.
The MD reaction carries reducing equivalents from glycolysis into the mitochondrial matrix as part of the malate-aspartate shuttle. [ Note: The NADH from glycolysis is oxidized as OAA is reduced to malate in the cytosol. Malate is transported from the cytosol into the mitochondrial matrix, where it is reoxidized to
OAA as NAD+ is reduced to NADH. No transporter exists to move NADH (or OAA) across the inner mitochondrial membrane.]
Tricarboxylic Acid Cycle 9.4
The delta Go for the conversion of malate to OAA is + 7.1 kcal/mol. What drives this reaction in the forward direction?
The endergonic conversion of malate to OAA ( delta G 0 of + 7.1 kcal/mol) is driven in the forward direction because it is coupled to the highly exergonic CS reaction
Tricarboxylic Acid Cycle 9.4
The addition of a small amount of malate or OAA to actively respiring muscle gives rise to a five-fold increase in O2 consumption. Why?
O2 consumption is a measure of the activity of aerobic pathways such as the TCA cycle. The addition of malate or OAA stimulates the TCA cycle, thereby increasing production of reducing equivalents for the ETC, the major O2 consumer.
Tricarboxylic Acid Cycle Summary 9.5
How many ATPs will be produced from the four reducing equivalents generated in one round of the TCA cycle, as shown?
Three NADHs and one FADH2 are generated in one round of the TCA cycle. Assuming a P/O ratio of 3 for NADH and 2 for FADH2,
11 ATPs will be generated by OXPHOS. Additionally, one GTP is
produced by substrate-level phosphorylation. GTP and ATP are interconverted by nucleoside diphosphate kinase. Therefore, a total of 12 ATP are produced by one round of the TCA cycle.
Tricarboxylic Acid Cycle Summary 9.5
What are the major TCA cycle regulation sites, and what are the effectors?
The major sites of cycle regulation (and the regulators) are CS (inhibited by citrate, its product), ICD (allosterically inhibited by NADH and ATP and activated by ADP and Ca2+ ), and the A-KGD
complex (allosterically inhibited by NADH and succinyl CoA and activated by Ca2+).
