Robert K. Preston

Absolute configuration of the isomer of fluorocitrate that inhibits aconitase

Abstract The absolute configuration of the isomer of fluorocitrate that is a strong inhibitor and activator of aconitase has been determined to be 2 R , 3 R , as shown in the Fischer and perspective diagrams below. Fischer diagram Perspective diagram This is the isomer described by Dummel and Kun (1969, J. Biol. Chem. 244 , 2966) and defined as (−)- erythro -fluorocitrate by them. We report here structural studies of the mirror image (enantiomer) of this isomer. Crystals of the complex of the diethyl ester of (+)- erythro -fluorocitrate with (−)-methylbenzylamine were studied by X-ray diffraction techniques. Since the absolute configuration of (−)-methylbenzylamine has already been determined experimentally, the absolute configuration of the (+)-erythro isomer of fluorocitrate is thereby established. This isomer is shown to be noninhibitory with the enzyme aconitase, while its racemate is a powerful inhibitor. Thus it is proved that the absolute configuration of the isomer of fluorocitrate that is formed from fluoroacetyl-CoA by the enzyme citrate synthase, and that inhibits aconitase, is the 2 R , 3 R , isomer.

Carboxylic-phosphoric mixed anhydrides isosteric with AMP and ATP as reagents for enzymic AMP and ATP sites.

Summary A carboxylic-phosphoric mixed anhydride was synthesized which differs from adenosine 5′-triphosphate (ATP) by replacement of the 5′ methylene group of ATP by a carbonyl group. This compound did not inhibit rabbit adenylate kinase, but it rapidly inactivated rabbit pyruvate kinase; the effect was prevented by ATP, ADP and phosphoenolpyruvate and therefore appears to be ATP-site-directed. The analogous anhydride isosteric with adenosine 5′-phosphate (AMP) rapidly inactivated rabbit AMP aminohydrolase; inactivation was prevented by an equimolar level of AMP. These findings, together with previous studies with this AMP analog, suggest that these anhydrides are powerful and potentially useful reagents for adenine nucleotide binding sites of enzymes.