Influence of Temperature and pH on the Proteolytic Activity of Rennet Extract

Abstract

The influence of incubation temperature and pH on the proteolytic activity of ren- net extract and crystalline rennin was in- vestigated. The major results revealed that changes in the electrophoretic pattern of casein gave a much better evaluation of the proteolytic activity of rennin than did changes in nonprotein nitrogen. With ca- sein as substrate and using changes in elec- trophoretie patterns as the criterion, the pH optimum for rennin protcolysis was about 5.8. Changes in nonprotein nitrogen indicated a much lower ptI optimum. The proteoIysis products were pH-dependent and similar results were obtained with both cls~stalline rennin and rennet extract. The nature of proteolysis products was not tem- perature-dependent but at low temperature fl-casein was more susceptible to proteolysis than asl-casein , whereas the reverse applied at higher temperature. Rennin is a weakly proteolytic enzyme (2, 3) and it is reasonable to assume that it plays some part in the degradation of proteins of cheese during ripening. The breakdown of proteins of cheese and of a- and fi-caseins in solution was investigated by Lindqvist and Storgards (16-19), using free-boundary elec- trophoresis. They concluded that both in cheese and in solution fl-casein was hydrolyzed more quickly than a-casein, but both underwent ex- tensive proteolysis. Ledford, O'Sullivan, and Nath (14), using polyacrylamide gel electro- phoresis, demonstrated that in Cheddar cheese the a~l-casein was specifically degraded, leaving the fl-easein essentially unaltered. In other cheese varieties, notably those surface ripened, fl-easeLn was also degraded, probably by the action of proteases secreted by ripening microorganisms. Further work by Ledford, Chen, and Nath (15), in which solutions of sodium caseinate and of sodium a~l- and fi- easeinates were subjected to the action of ren- net, confirmed that asl-casein was more suscep- tible to proteolysis by rennet than was fl- casein. These workers also found that casein