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Unformatted text preview: Lecture 9 Lecture 9: Citric Acid Cycle 16.1 Producing Acetyl-CoA- many sugars, fatty acids, amino acids enter the cycle by degrading into the acetyl group of acetyl-CoA - pyruvate is oxidized to acetyl-coA and 2 CO by pyruvate dehydrogenase complex (PDH) - Chemical intermediates are bound to the enzyme as substrate is converted to product- PDH performs an oxidative decarboxylation where the carboxyl group is removed as 2 CO , electrons transferred to NAD+ to form NADH- PDH complex is irreversible and cannot reattach radioactively labeled 2 CO - PDH requires 3 enzymes and 5 coenzymes/prosthetic groups that contain vitamines: TTP (thiamine), FAD (riboflavin), CoA (panthothenate), NAD, lipoate- Coenzyme A has a reactive thiol group; acyl groups are linked to thiol groups forming thioesters (the acyl group attached to coenzyme A is activated for group transfer - Lipoate has 2 thiol groups that undergo reversible oxidation to disulfide bond, one SH can carry an acetyl group - Lipoic acid is in amide linkage with a Lys residue More about PDH contains 3 enzymes: pyruvate dehydrogenase (E 1 ), dihydrolipoyl transacetylase (E 2 ) and dihydrolipoyl dehydrogenase (E 3 ) E1 active site as bound TPP, E2 has covalently bound lipoyl group, E3 has FAD and NAD; 2 regulatory proteins: protein kinease and phosphoprotein phosphatase The organization of the PDH complex is very similar to the enzyme complexes that catalyze the oxidation of -ketoglutarate and branched chains of -keto acids Substrate Channeling- intermediates never leave the enzyme surface- in step 1) C1 of pyruvate is release as 2 CO and C2 (oxidation state of aldehyde) is attached to TPP as a hydroxyethyl group- This first step is slowest and limits rate of overall reaction; important for substrate specificity - Step 2) hydroxyethyl group is oxidized to level of carboxylic acid (acetate) - 2 electrons removed reduce disulfide bond of lipoyl group to two thiol groups- Acetyl group is first esterified to one of the SH groups, then transesterified to form acetyl-coA- Steps 4/5 regenerate the oxidized (disulfide) form of the lipoyl group 1 of 8 Lecture 9- Electrons removed from the hydroxyethyl group derived from pyruvate pass through FAD to NAD+- All these enzymes are clustered so the intermediates never leave the complex, local concentration of substrate E2 is kept very hight 16.2 The Reactions of the Citric Acid Cycle each turn, one acetyl-group (2 carbons) enters as acetyl-CoA and 2 molecules of 2 CO leave; one molecule of oxaloacetate is regenerated Oxaloacetate is recycled and present in cells in low concentrations...
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