Chapter 19

The malate synthase reaction, which produces malate from acetylCoA and glyoxylate in the glyoxylate pathway, involves chemistry similar to the citrate synthase reaction. Write a mechanism for the malate synthase reaction and explain the role of CoA in this reaction.

In most cells, fatty acids are synthesized from acetate units in the cytosol. However, the primary source of acetate units is the TCA cycle in mitochondria, and acetate cannot be transported directly from the mitochondria to the cytosol. Cells solve this problem by exporting citrate from the mitochondria and then converting citrate to acetate and oxaloacetate. Then, because cells cannot transport oxaloacetate into mitochondria directly, they must convert it to malate or pyruvate, both of which can be taken up by mitochondria. Draw a complete pathway for citrate export, conversion of citrate to malate and pyruvate, and import of malate and pyruvate by mitochondria. a. Which of the reactions in this cycle might require energy input? b. What would be the most likely source of this energy? c. Do you recognize any of the enzyme reactions in this cycle? d. What coenzymes might be required to run this cycle?

Glycolysis, the pyruvate dehydrogenase reaction, and the TCA cycle result in complete oxidation of a molecule of glucose to \(\mathrm{CO}_{2}\). Review the calculation of oxidation numbers for individual atoms in any molecule, and then calculate the oxidation numbers of the carbons of glucose, pyruvate, the acetyl carbons of acetyl-CoA, and the metabolites of the TCA cycle to convince yourself that complete oxidation of glucose involves removal of 24 electrons and that each acetyl-CoA through the TCA cycle gives up 8 electrons.

Aconitase is rapidly inactivated by \(2 R, 3 R\) -fluorocitrate, which is produced from fluoroacetate in the citrate synthase reaction. Interestingly, inactivation by fluorocitrate is accompanied by stoichiometric release of fluoride ion (i.e., one F-ion is lost per aconitase active site \() .\) This observation is consistent with "mechanism-based inactivation" of aconitase by fluorocitrate. Suggest a mechanism for this inactivation, based on formation of 4 -hydroxy-trans-aconitate, which remains tightly bound at the active site. To assess your answer, consult this reference: Lauble, H., Kennedy, M., et al., 1996. The reaction of fluorocitrate with aconitase and the crystal structure of the enzyme-inhibitor complex. Proceedings of the National Academy of Sciences \(93: 13699-13703\)