EFFECT OF pH UPON PROTEOLYSIX BY PAPAIN” BY SAM R. HOOVER

The existence of pH optima for proteolysis was early established, primarily through measurements of the initial rate of reaction as a function of pH. When synthetic substrates for proteolytic enzymes were discovered, it was found that the rate of digestion depended upon the amino acids of the peptide and their arrangement in the chain. We have investigated the effect of pH upon the hydrolysis of peptide bonds of different specific rates of digestion in an endeavor to find out whether a differential effect of pH upon the extent of enzymic hydrolysis of proteins could be demonstrated. The evidence available in the literature was indirect, but was consistent with such a pH effect. The optimum pH for the initial digestion of different proteins by papain varies, being about 5 for the digestion of gelatin and 7 for casein (1) and egg albumin (2). Lineweaver and Schmimmer (1) found a similar PI-I-activity function for the action of both crystalline and commercial papain on casein. The initial rate of digestion at pH 5 and 9 was about 60 per cent of that at pH 6.5 to 7.0. This was true both when the casein was in suspension in the isoelectric region and when it was held in solution by urea. Rergmann and coworkers used pH 5 throughout in their study of the digestion of synthetic substrates by pa,pain. Rocha e Silva and Andrade (3) obtained evidence of two pH opt’ima, about 5 and 7, for the action of papain upon benzoylargininamide. Pepsin digests synthetic substrates less rapidly at it,s normal pH optimum of about 2 than at pH 4.2 (4, 5). The latter pH was used successfully by Pope for t’he purification of diphtheria antitoxin by pepsin (6). We have investigated the effect of pH upon the course of proteolysis by papain, using two substrate systems: (a) the synthetic substrates of Bergmann and coworkers, in which a first order reaction can be demonstrated and (b) casein. The rate and extent of proteolysis of casein at pH 7, 5, and 2.5 were determined. An optimum pH of about 5 for the “peptidase” action of papain was demonstrated in both systems. The extent of hydrolysis of casein is far greater at pH 5 than at the optimum for initial hydrolysis, about pH 7.