Photoaffinity labeling of Klebsiella aerogenes citrate lyase by p-azidobenzoyl-CoA

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Abstract
P-Azidobenzoyl CoA functions as a linear competitive inhibitor for (3S)-citryl-CoA in the citrylCoA oxalacetate-lyase reaction catalyzed by the K. aerogenes deacetylcitrate lyase complex (Ki = 80 .mu.M; (3S)-citryl-CoA Km = 67 .mu.M). Inactivation is irreversible on photolysis of p-azidobenzoyl-CoA in the presence of the deacetylcitrate lyase complex. Mg2+ is not required for the inactivation. Inactivation is blocked by (3S)-citryl-CoA in the presence of EDTA. p-Azidobenzoyl-CoA has no effect on the acetyl-CoA:citrate CoA transferase activity of both the deacetylcitrate lyase complex and its isolated transferase subunit. The stoichiometry of the CoA ester binding was investigated by the use of p-azido[14C]benzoyl-CoA as a photoaffinity reagent. The labeling is exclusively on the lyase .beta. subunit of the citrate lyase (EC 4.1.3.6) complex.