3-fluoro-3-deoxycitrate: a probe for mechanistic study of citrate-utilizing enzymes

The interaction of a novel fluorinated analogue of citrate, 3-fluoro-3-deoxycitrate (3-fluorocitrate), with the four known citrate-processing enzymes is described in this report. Three of the citrate-processing enzymes, citrate synthase, ATP citrate lyase, and citrate lyase, catalyze reversible aldol-type condensations. The fate of 3-fluorocitrate with each enzyme is uniquely related to their mechanisms of action. For citrate synthase, 3-fluorocitrate is a competitive inhibitor. 3-Fluorocitrate is a substrate for the carboxylate activation half-reaction catalyzed by ATP citrate lyase and induces a net ATPase action during conversion to 3-fluorocitryl-S-coenzyme A. Because of the unusual mechanism of citrate cleavage catalyzed by bacterial citrate lyase, 3-fluorocitrate is a mechanism-based inhibitor, acting at two points during turnover of the acetyl enzyme. The fourth citrate-processing enzyme, aconitase, does turn over 3-fluorocitrate catalytically. This enzyme, catalyzing a dehydration and rehydration of citrate, also catalyzes the elimination of HF from 3-fluorocitrate, yielding cis-aconitate and fluoride.