Enzymatic Synthesis of Acetono-CoA, A Competitive Inhibitor of Acetyl-CoA with Citrate Synthase
The synthetic acetyl-CoA analog, acetono-CoA (acetonyldethiocoenzyme A), in which the sulfur atom has been replaced by a methylene group, has been previously synthesized and found to be a competitive inhibitor (KI= 13.2 µM) of acetyl-CoA with citrate synthase. To determine whether the acetono-group would interfere with CoA biosynthesis, a crude preparation of the bifunctional enzyme complex which converts ATP and pantetheine 4’-phosphate to CoA was investigated. Incubation mixtures of the crude complex, ATP, and acetonyldethio-pantetheine 4’-phosphate were prepared. Products and reactants were isolated by DEAE-cellulose column chromatography and peaks were identified by TLC. Enzymatic synthesis of acetono-CoA was verified by establishing its KI (25 µM) in the citrate synthase reaction.
pantetheine 4’-phosphate + ATP ---> dephospho-CoA + PP
dephospho-CoA + ATP ---> CoA + ADP
is catalyzed by the bifunctional coenzyme A synthesizing enzyme complex, 1) pantetheine-phosphate adenyltransferase (EC 126.96.36.199) and 2) dephospho- CoA kinase (EC 188.8.131.52). We investigated the ability of the crude enzyme complex to convert ATP and the pantetheine-phosphate analog, acetonyldethiopantetheine 4’- phosphate to acetono-CoA.
KeywordsEnzymatic Synthesis Crude Enzyme Preparation Acetyl Phosphate Oxaloacetic Acid Methylene Analog
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- 1.Stewart, C.J. and Wieland, T. (1978), Liebigs Ann. Chem. 57–65Google Scholar
- 4.Bandurski, R.S. and Axelrod, B. (1951) J. Biol. Chem. 193, 405–410Google Scholar
- 5.Hoagland, M.B. and Novelli, G.D. (1954) J. Biol. Chem. 207, 767–773Google Scholar
- 6.Abiko, Y. (1971) Methods Enzymol. 18A, 358–365Google Scholar
- 7.Abiko, Y., Suzuki, T., Shimizu, M. (1967) J. Biochem. 61, 10–17Google Scholar