PowerPoint PresentationSlide 2Slide 3Pathway Overview The eight reactions of the citrate cycle oxidize acetyl-CoA to generate 2 CO2, and in the process, reduce 3 NAD+ and 1 FAD. In addition, 1 GTP is produced by substrate level phosphorylation which is converted to ATP by nucleotide kinase.Slide 5Slide 6Hans Krebs Elucidated the Citrate CyclePathway QuestionsSlide 9Slide 10Slide 11Slide 12The complete oxidation of glucose to CO2 and H2O is summarized by the reaction:Eight Reactions of the Citrate CycleReaction 1: Condensation of oxaloacetate and acetyl-CoA by citrate synthase to form citrateReaction 2: Isomerization of citrate by aconitase to form isocitrateAconitase is one of the targets of fluorocitrateReaction 3: Oxidative decarboxylation of isocitrate by isocitrate dehydrogenase to form α-ketoglutarate, CO2 and NADHReaction 4: Oxidative decarboxylation of by α-ketoglutarate dehydrogenase to form succinyl-CoA, CO2 and NADHReaction 5: Conversion of succinyl-CoA to succinate by succinyl-CoA synthetase in a substrate level phosphorylation reaction that generates GTPReaction 6: Oxidation of succinate by succinate dehydrogenase to form fumarateSlide 22Reaction 7: Hydration of fumarate by fumarase to form malateReaction 8: Oxidation of malate by malate dehydrogenase to form oxaloacetateBioenergetics of the citrate cycleSlide 26Radioactive 14C-acetyl CoASlide 28Slide 29Citrate Cycle:Energy conversion through redox reactions, a cycle with eight renewable reactants, and regulation Bioc 460 Spring 2008 - Lecture 28 (Miesfeld)If the citrate cycle is an engine, what is the fuel, the energy output, and the exhaust?Compound 1080 contains fluoracetate, a poison that blocks the activity of aconitaseThe Citrate Cycle is spinning out of control!•The primary function of the citrate cycle is to convert energy available from the oxidization acetyl-CoA into 3 moles of NADH, 1 mole of FADH2 and 1 mole of GTP during each turn of the cycle.•The citrate cycle is a "metabolic engine" in which all eight of the cycle intermediates are continually replenished to maintain a smooth-running energy conversion process. The fuel for this metabolic engine is acetyl-CoA, the exhaust is two molecules of CO2, and the energy output is redox energy used in the electron transport system for ATP synthesis.•Oxaloacetate is both the product of reaction 8, and the reactant for reaction 1, which means that flux through the pathway is continuously monitored by resetting the level of available substrate after each turn of the cycle. Two other regulatory mechanisms at play in the citrate cycle are product inhibition and feedback control of key enzymes.Key Concepts in Citrate CycleThe citrate cycle is considered the "hub" of cellular metabolism because it not only links the oxidation of metabolic fuels (carbohydrate, fatty acids and proteins) to ATP synthesis, but it also provides shared metabolites for numerous other metabolic pathways.Pathway OverviewThe eight reactions of the citrate cycle oxidize acetyl-CoA to generate 2 CO2, and in the process, reduce 3 NAD+ and 1 FAD. In addition, 1 GTP is produced by substrate level phosphorylation which is converted to ATP by nucleotide kinase.All of the enzymes in the citrate cycle, electron transport chain and oxidative phosphorylation reside in the mitochondrial matrix where pyruvate is converted to acetyl-CoA by the enzyme pyruvate dehydrogenase.The "currency exchange" for redox energy and ATP synthesis in the mitochondria electron transport chain is ~2.5 ATP/ NADH. Oxidation of 2 FADH2 molecules by the electron transport chain results in only ~3 molecules of ATP (~1.5 ATP/FADH2) because of differences in where these two coenzymes enter the electron transport chain. Based on this ATP currency exchange ratio, and the one substrate level phosphorylation reaction, each turn of the cycle produces ~10 ATP for every acetyl-CoA that is oxidized.Hans Krebs Elucidated the Citrate CycleHans Krebs, a biochemist who fled Nazi Germany for England in 1933, first described the citrate cycle in 1937. The citrate cycle is sometimes called the Krebs cycle, the citric acid cycle, or the tricarboxylic acid cycle, although we will refer to it as the citrate cycle because citrate is the first product of the pathway.The unprotonated form of citric acid is citrate which is the predominant species at physiological pH (the pKa values of the three carboxylate groups are 3.1, 4.7 and 6.4).What does the citrate cycle accomplish for the cell?–Transfers 8 electrons from acetyl-CoA to the coenzymes NAD+ and FAD to form 3 NADH and 1 FADH2 which are then re-oxidized by the electron transport chain to produce ATP by the process of oxidative phosphorylation.–Generates 2 CO2 as “waste products” and uses substrate level phosphorylation to generate 1 GTP which is converted to ATP by nucleoside diphosphate kinase.–Supplies metabolic intermediates for amino acid and porphyrin biosynthesis.Pathway QuestionsWhat is the overall net reaction of citrate cycle?Acetyl-CoA + 3 NAD+ + FAD + GDP + Pi + 2 H2O →CoA + 2 CO2 + 3 NADH + 2 H+ + FADH2 + GTPΔGº’ = -57.3 kJ/molPathway QuestionsIs the citrate cycle a favorable reaction? Of course it is, you are alive aren’t you!Pathway QuestionsWhat are the key regulated enzymes in citrate cycle?Pyruvate dehydrogenase – not a citrate cycle enzyme but it is critical to flux of acetyl-CoA through the cycle; this multisubunit enzyme complex requires five coenzymes, is activated by NAD+, CoA and Ca2+ (in muscle cells), and inhibited by acetyl-CoA, ATP and NADH.Citrate synthase – catalyzes the first reaction in the pathway and can be inhibited by citrate, succinyl-CoA, NADH and ATP; inhibition by ATP is reversed by ADP.Pathway QuestionsWhat are the key regulated enzymes in citrate cycle?Isocitrate dehydrogenase - catalyzes the oxidative decarboxylation of isocitrate by transferring two electrons to NAD+ to form NADH, and in the process, releasing CO2, it is activated by ADP and Ca2+ and inhibited by NADH and ATP.α-ketoglutarate dehydrogenase - functionally similar to pyruvate dehydrogenase in that it is a multisubunit complex, requires the same five coenzymes and catalyzes an oxidative decarboxylation reaction that produces CO2, NADH and succinyl-CoA; it is activated by Ca2+ and AMP and it is inhibited by NADH, succinyl-CoA and ATP.Pathway QuestionsWhat are examples of citrate cycle in real life?Citrate is produced commercially by fermentation methods using
View Full Document
Unlocking...