The kinetic mechanism of inhibition of human leukocyte elastase by MR889, a new cyclic thiolic compound

Abstract

The cyclic thiolic compound 2-[3-thiophencarboxythio]- N-[dihydro-2(3 H)-thiophenone1-3-il]-propionamide (MR889) was investigated as inhibitor of endopeptidases. The activity of bovine pancreatic α-chymotrypsin, human leukocyte cathepsin G and rabbit liver cathepsin B was not affected by MR889, whereas porcine pancreatic elastase and human leukocyte elastase were inhibited. The kinetic mechanism of inhibition of human leukocyte elastase was of the reversible, slow-binding, fully competitive type. The rate constants for complex formation between MR889 and leukocyte elastase, determined by pre-steady-state kinetic analysis in the presence of a tetrapeptide substrate at 37° and pH 7.40, were k on = 2363 ± 15 M −1 sec −1, k off = 3.01 ± 0.34 × 10 −3 sec −1. The inhibition equilibrium constant was k i = k off k on = 1.27 ± 0.15μM . K i , calculated from steady-state kinetic experiments, was 1.38 μM. MR889 also inhibited the elastolytic activity of leukocyte elastase, as determined with insoluble elastin as the substrate.