Yasuhiro Hosokawa

Structural specificity of coenzyme a for phosphotransacetylase

Abstract The structural requirements for the binding of CoA to phosphotransacetylase from two sources, Escherichia coli B and Clostridium kluyveri, were investigated by kinetic analysis of the effects of various CoA analogues. Results are expressed in terms of type of inhibition with respect to CoA, Ki value, and coenzyme activity. Analogues lacking the cysteamine, the aletheine and the pantetheine moieties, as well as adenosine 3′-phosphate 5′-pyrophosphate, which corresponds to the nucleotide moiety of the coenzyme, were found to compete with CoA. Moreover, with this nucleotide the enzyme could acetylate pantetheine with acetyl phosphate in the absence of CoA. Desulfo-CoA, a potent competitive inhibitor of CoA, inhibited the acetylation of pantetheine. These results suggest that this nucleotide interacts specifically with a CoA-site of the enzyme, indicating that the nucleotide moiety of CoA plays an essential role in determining the specificity of this coenzyme for phosphotransacetylase. This was supported by the finding that analogues modified on the nucleotide moiety scarcely interact with the enzyme. CoA analogues with modified cysteamine and pantothenic acid moieties also competed with CoA, but their affinities for the enzyme were greatly decreased. These results, together with the above observations, suggest that the pantetheine moiety may not substantially be involved in the specificity of CoA for the enzyme, but it contributes significantly to the strength of binding of CoA to phosphotransacetylase.

Effect of adenine nucleotide moiety of coenzyme A on phosphotransacetylase

Abstract Structural analogues of CoA lacking in aletheine moiety as well as in pantetheine moiety were found to compete with CoA in the phosphotransacetylase reaction. Moreover, the latter compound has induced an ability of the enzyme to acetylate pantetheine in the absence of CoA, which was inhibited by desulfo-CoA. These results suggest that adenine nucleotide moiety of CoA plays an essential role in being recognized as CoA by phosphotransacetylase.