Flashcards in Lecture 34 Deck (31)
What are the three major steps of catabolism leading to the TCA cycle?
1) Hydrolysis of complex molecules (proteins, polysaccharides, & lipids) to their component building blocks (amino acids, monosaccharides & glycerol + fatty acids)
2) Conversion of building blocks to acetyl CoA
3) Oxidation of acetyl CoA; oxidative phosphorylation
What are the steps to the mechanism of action of the pyruvate dehydrogenase complex?
1) Pyruvate is decarboxylated to form a hydroxyethyl derivative bound to the reactive carbon of thiamine pyrophosphate (TPP), the coenzyme of pyruvate dehydrogenase
2) The hydroxyethyl intermediate is oxidized by transfer to the disulfide form of lipoic acid covalently bound to dihydrolipoyl transacetylase
3) The acetyl group, bound as a thioester to the side chain of lipoic acid, is transferred to CoA
4) The sulfhydryl form of lipoic acid is oxidized by FAD-dependent dihydrolipoyl dehydrogenase, leading to the regeneration of oxidized lipoic acid
5) The reduced flavoprotein is reoxidized to FAD by dihydrolipoyl dehydrogenase as NAD+ is reduced
How does the pyruvate dehydrogenase complex become inactivated?
Phosphorylation by a protein kinase (converts ATP --> ADP)
What are cofactors that stimulate the function of the protein kinase to phosphorylate the pyruvate dehydrogenase complex?
2) Acetyl CoA
What are cofactors that inhibit the function of the protein kinase, so that it does not phosphorylate the pyruvate dehydrogenase complex?
How does the pyruvate dehydrogenase complex become activated?
Phospho-protein phosphatase removes the phosphorylation of pyruvate dehydrogenase complex with the use of H2O
What stimulates the function of Phospho-protein phosphatase?
What directly reduces the functionality of the active pyruvate dehydrogenase complex?
2) Acetyl CoA
What two main reactants combine to form the 6 carbon citrate in the citric acid cycle?
1) Acetyl-CoA (2C)
2) Oxaloacetate (4C)
What carbon compound is produced by the TCA?
Oxaloacetate (used as a reactant)
What are the first three steps of TCA?
1) Acetyl CoA + Oxaloacetate + H2O --> CoA + Citrate (catalyzed by citrate synthase)
2) Citrate --> Isocitrate (catalyzed by aconitase, which allows the movement of the hydroxyl group)
3) Isocitrate + NAD+ --> alpha ketoglutarate + CO2 + NADH + H+ (catalyzed by isocitrate dehydrogenase)
What are steps 4-7 of the TCA?
1) alpha ketoglutarate + NAD+ + CoA --> CO2 + NADH + H+ + Succinyl CoA (catalyzed by alpha-ketoglutarate dehyrogenase complex)
2) Succinyl CoA + GDP + Pi --> GTP + CoA + Succinate (catalyzed by Succinyl CoA thiokinase)
3) Succinate + FAD --> Fumarate + FADH2 (catalyzed by succinate dehydrogenase)
4) Fumarate + H2O --> L-Malate (catalyzed by fumarase)
What is the last step of the TCA?
L-Malate + NAD+ --> Oxaloacetate + NADH + H+ (catalyzed by Malate Dehydrogenase)
What are the three components of the pyruvate dehydrogenase complex?
1) E1 - Pyruvate Dehydrogenase (contains thiamine pyrophosphate (TPP) bound to it)
2) E2 - Dihydrolipoyl transacetylase (lipoic acid transacetylase)
3) E3 - Dihydrolipoyl dehydrogenase
What are the functions of E1, E2, & E3 in the pyruvate dehydrogenase complex?
1) E1 - Removes carboxyl group from Pyruvate to form hydroxyethyl TPP and release CO2
2) E2 - Forms a high energy thioester bond between sulfur on lipoic acid and acetyl group (hydroxyethyl group) from TPP
3) CoA-SH forms high energy bond between its sulfur group and acetyl group from lipoic acid (transacetylase transfers acetyl group) to form Acetyl CoA
4) Must reform disulfide bridge in lipoic acid by reducing FAD to FADH2 (from E3)
5) Must reform FAD by using NAD+ to oxidize FADH2
What are the 3 dehydrogenases in the TCA cycle that reduce NAD+ to NADH + H+?
1) Isocitrate dehydrogenase
2) Alpha-ketoglutarate dehydrogenase complex
3) Malate dehydrogenase
All of the enzymes used in the TCA cycle function in the ____. However, the enzyme _____ also functions in the ____.
2) Malate dehydrogenase
What is a short summary of the TCA cycle?
1) Two carbon atoms enter the cycle
2) Two molecules of CO2 are released during the cycle
3) Substrate-level phosphorylation occurs when Succinyl CoA is split to form ATP + GTP
4) Four reduced cofactor molecules per acetyl CoA are oxidized to CO2 (3 NADH + 1 FADH2)
What are inhibitors of the pyruvate dehydrogenase complex and the TCA cycle?
1) Enzyme: PyruvateDH
Inhibitors: ATP, NADH, & Acetyl CoA
2) Enzyme: Citrate synthase
3) Enzyme: IsocitrateDH
INhibitors: ATP & NADH
4) Enzyme: alpha ketoglutarate DH
Inhibitor: ATP & NADH
What are activators of the TCA cycle?
1) Enzyme: Isocitrate DH
Can the intermediate compounds formed in glycolysis and the TCA cycle be used in other reactions?
Yes, they can be used in many metabolic pathways
What is a result of competing reactions using intermediates from glycolysis and the TCA cycle?
This diversion of intermediates creates a need for reactions that will maintain the level of oxaloacetate
What is anaplerosis?
1) It is one of the most important reactions for the production of oxaloacetate catalyzed by pyruvate carboxylase
2) High levels of pyruvate carboxylase for gluconeogenesis occur in the liver and kidney, but other cells have lower activities of pyruvate carboxylase to maintain the concentration of oxaloacetate
What are mutations of TCA cycle genes in cancer?
1) Mutations in isocitrate dehydrogenase 1 and 2 result in the formation of 2-hydroxyglutarate (2HG) instead of alpha-ketoglutarate. 2HG is a competitive inhibitor of alpha-ketoglutarate-dependent dioxygenases. Dioxygenases have an important role in demethylation reactions for histones and DNA causing hypermethylation in glioma and acute myelocytic leukemia.
2) Low activities of fumarate hydratase (fumarase) drives a metabolic shift to aerobic glycolysis in some kidney tumors and thereby enhances the Warburg effect in which aerobic glycolysis tends to be increased in cancer cells
What is the Warburg effect?
The Warburg effect is the observation that most cancer cells predominantly produce energy by a high rate of glycolysis followed by lactic acid fermentation in the cytosol, rather than by a comparatively low rate of glycolysis followed by oxidation of pyruvate in mitochondria as in most normal cells (even in the presence of oxygen)
Why must electrons be shuttled between the cytosol and mitochondria?
The inner mitochonrial membrane is impermeable to NADH, therefore electrons from NADH in the cytsol are transferred by electron shuttles
What is the glycerol phosphate shuttle?
In the glycerol phosphate shuttle, In the glycerol phosphate shuttle, NADH in the cytosol is used to reduce dihydoxyacetone phosphate in the reaction catalyzed by cytosolic glycerol 3-phosphate dehydrogenase. A glycerol phosphate oxidase in the inner mitochondrial membrane catalyzes the transfer of electrons from dihydroxyacetone phosphate to FAD. Reduced FAD is oxidized by Complex II in the mitochondrial electron transport chain
What is the malate-aspartate shuttle?
The malate-aspartate shuttle is an alternative mechanism for the
transfer of electrons into mitochondria.
What is a summary of the glycerol phosphate shuttle?
1) DHAP + NADH + H+ --> NAD+ + Glycerol 3-phosphate (occurs in cytosol)
2) Glycerol 3-phosphate + FAD --> FADH2 + DHAP (occurs in inner mitochondrial membrane; FADH2 can now be used in the ETC)
Under anaerobic conditions, the TCA cycle is limited by low levels of?