The specific esterase activity of trypsin.

The discovery of relatively simple peptide derivatives as substrates for proteolytic enzymes has introduced a new approach to the problem of the specificity of this group of hydrolytic enzymes. Although crystalline trypsin is known to catalyze the hydrolysis of internal peptide bonds of proteins, its action toward internal peptide bondsof synthetic substrates has been demonstrated for only one case (1). Since all other synthetic substrates for trypsin, described in the extensive work of Bergmann and coworkers (2), are amides of lysine and arginine derivatives and are split enzymatically into ammonia and the corresponding amino acid derivatives, the action of trypsin toward these substrates may be more adequately described as an amidase activity. In the course of an investigation of the inhibition of crystalline pancreatic proteolytic enzymes by specific low molecular weight compounds the discovery was made that crystalline trypsin is likewise a powerful catalyst for the hydrolysis of certain amino acid esters. The most specific substrates of this type are cY-benzoyl-L-arginine methyl ester (BAME) and cu-toluenesulfonyl-L-arginine methyl ester (TSAME). It was also found that, contrary to previous reports (3), trypsin catalyzes the amide hydrolysis of cr-toluenesulfonyl-L-argininamide and does so even more rapidly than it catalyzes the hydrolysis of the classical substrate, cr-benzoyl+argininamide. The structural specificity of these esters for trypsin is approximately analogous to that of the corresponding amides, although a certain measure of cross-reactivity of one of these (BAME) with chymotrypsin exists. Both the e&erase and amidase activities are apparently mediated by the same active surface configurations of the enzyme, as is evidenced by studies of various modes of partial and complete enzyme inactivation. The results of these kinetic studies are presented in this paper.