1. Ferchmin 2016 I ndex: 1.Its multifunctional role in the metabolism 2.Pyruvate dehydrogenase complex, structure, cofactors, mechanism, regulation. 3.TCA or Kreb’s cycle, function and regulation. 4.“Accounting” of ATP synthesized, from glycolysis to respiraory chain. 5.Murphy’s Law and the missing 10 molecules of ATP per glucose. 6.How to avoid losing your carbons during the exam. 1

2. Steps shown and enzymes involved. 1 and 2) Pyruvate dehydrogenase (not decarboxylase). 3 and 4) Dihydrolipoyl transacetylase. 5) Dihydrolipoyl dehydrogenase Fully reduced lipoic How come NAD + +H transfers H to FAD? The –SH reduces the other –S- of lipoic acid 2

3. Regulation of the PDC by allosteric and covalent modulation The enzymes involved CoenzymesDescriptionActivatorsInhibitors 1) Pyruvate dehydrogenase (E 1 ) Thiamine pyrophosphate (Vitamin B1) Decarboxylation of pyruvate, E 1 α and E 1 β, phosphorylation site of PDC. 30 subunits AMP energy GTP energy 2) Lipoate transacetylase (E 2 ) Lipoic acidDehydrogenation of hydroxyethyl and transfer of acetyl to CoA. 60 subunits CoA TCA substrate CoA~Acetyl TCA substrate 3) Dihydrolipoyl dehydrogenase (E 3 ) NAD + and FAD (Niacin & B2) Regeneration of the lipoate and reduction of NAD + to NADH. 10 subunits NAD + Reducing power NADH; Reducing power The regulatory subunits of the PDC are intrinsic to the complex 1) Pyruvate dehydrogenase kinase inactivates the PDC Complex by phosphorylation of three serines on E 1 α. PRODUCTS ATP Acetyl CoA NADH SUBSTRATES ADP; pyruvate; CoA; NAD + 2) Pyruvate dehydrogenase phosphatase removes the inhibition imposed by the phosphorylation through hydrolysis of the P i. Ca 2+ insulin NADH Ca 2+ in muscle & insulin in liver Insulin (means abundance of glucose) disinhibits the PDC and reroutes pyruvate from gluconeogenesis to lipogenesis 3

4. Stoichiometry of the Krebs Cycle: CH3-CO-CoA + 3 NAD+ FAD + GDP + Pi + 2 H 2 O  2 CO 2 + 3 NADH + FADH 2 + GTP + 2 H + + CoA ΔG’°=1.1 kcal/mole (sluggish) but citrate leaves the mito. and serves as substrate for lipid synthesis Ac~CoA never leaves mito to favor TCA Succinyl~CoA accumulates in mitosol and serves as feedback inhibitor of citrate synthase, donor of CoA~ to activate ketone bodies and fatty acids and is also a precursor of porphyrines. Isocitrate dehydrogenase is the key enzyme (committed step). Strictly dependent on the ration of ADP/ATP and NAD + /NADH. Makes TCA aerobic by the “substrate control”. Odd carbon number fatty acids enter here and contribute to gluconeogenesis exiting the TCA as malate. Availability of oxaloacetate (OAA) is one of the main limiting steps of TCA. The [OAA] is 1/10 of the other intermediates of TCA. Remember pyruvate carboxylase is anaplerotic. Why Acetyl- CoA activates pyruvate carboxylase? Do you remember from glycolysis that the active metabolite (glyceraldehyde) is often kept in short supply? Succinate dehydrogenase is part of the mito. membrane and the respiratory chain Malate leaves the mitosol and goes into gluconeogenesis Very similar to PDC but has no intrinsic protein kinases & phosphatases. Otherwise has ~ the same regulation The green arrow indicates that the equilibrium is displaced towards malate. 4

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6. The yield of ATP per NADH+H + depends on the shuttle used. 6

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