Citation

Garcia-Alles LF, Erni B. Eur. J. Biochem. 2002; 269(13): 3226-3236.

Copyright

(Copyright © 2002, Blackwell Science Ltd. on behalf of the Federation of European Biochemical Societies)

DOI

10.1046/j.1432-1033.2002.02995.x

PMID

12084063

Abstract

The synthesis of 10 new phosphoenolpyruvate (PEP) analogues with modifications in the phosphate and the carboxylate function is described. Included are two potential irreversible inhibitors of PEP-utilizing enzymes. One incorporates a reactive chloromethylphosphonate function replacing the phosphate group of PEP. The second contains a chloromethyl group substituting for the carboxylate function of PEP. An improved procedure for the preparation of the known (Z)- and (E)-3-chloro-PEP is also given. The isomers were obtained as a 4 : 1 mixture, resolved by anion-exchange chromatography after the last reaction step. The stereochemistry of the two isomers was unequivocally assigned from the (3)J(H-C) coupling constants between the carboxylate carbons and the vinyl protons. All of these and other known PEP-analogues were tested as reversible and irreversible inhibitors of Mg2+- and Mn2+- activated PEP-utilizing enzymes: enzyme I of the phosphoenolpyruvate:sugar phosphotransferase system (PTS), pyruvate kinase, PEP carboxylase and enolase. Without exception, the most potent inhibitors were those with substitution of a vinyl proton. Modification of the phosphate and the carboxylate groups resulted in less effective compounds. Enzyme I was the least tolerant to such modifications. Among the carboxylate-modified analogues, only those replaced by a negatively charged group inhibited pyruvate kinase and enolase. Remarkably, the activity of PEP carboxylase was stimulated by derivatives with neutral groups at this position in the presence of Mg2+, but not with Mn2+. For the irreversible inhibition of these enzymes, (Z)-3-Cl-PEP was found to be a very fast-acting and efficient suicide inhibitor of enzyme I (t(1/2) = 0.7 min).

Language: en

Keywords

Biochemistry; Drug Evaluation, Preclinical; Enzyme Activation; Enzyme Inhibitors; Isomerism; Phosphoenolpyruvate; Phosphoenolpyruvate Carboxylase; Phosphoenolpyruvate Sugar Phosphotransferase System; Phosphopyruvate Hydratase; Phosphotransferases (Nitrogenous Group Acceptor); Pyruvate Kinase; Structure-Activity Relationship