Kinetic Studies on the Interactions between Native, Acetylated and Succinylated Trypsin and Natural Proteinase Inhibitors

Abstract

It is now well established that the amino acid residues, which participate in the active site of trypsin, are located in the contact area between the enzyme and its natural proteinase inhibitors. This has been shown by several lines of experimental evidence: a) the inhibition of trypsin is competitive [1]; b) trypsin in which the active site Ser 183 is modified with diisopropylfluorophosphate (DFP) does not bind any longer the inhibitors [1, 2]; c) trypsin in which the active site His 46 is modified either by photooxidation [2, 3] or by tosyllysylchloro-methylketone (TLCK) [2], or by bromoacetone [4] does not bind some of the inhibitors; d) the pH dependence of the association rate constant between trypsin and the pancreatic [5, 6] or soybean Kunitz [7], or Kazal inhibitors [8] indicates that His 46 is involved in the association process; e) finally, the recent reports on the threedimensional structure of the trypsin-pancreatic Kunitz inhibitor complex [9] and the trypsin-soybean Kunitz inhibitor complex [10] confirm the kinetic and chemical studies.

Supported by grants from the Centre National de la Recherche Scientifique (Equipe de Recherche N 142), Délégation Générale à la Recherche Scientifique et Technique, contract number 71.3001 and Foundation pour la Recherche Médicale Française.