Ribonuclease E, an Intermediate in the Degradation of Ribonuclease by Porcine Elastase.

Bovine pancreatic ribonuclease has been studied as the substrate of a number of proteolytic enzymes. In the native state it is completely resistant to trypsin and chymotrypsin (1, 2), but denaturation by chemical (3) or physical (4, 5) means induces susceptibility to these enzymes. Ribonuclease is! however, susceptible to attack by a number of other proteases. Subtilisin has been shown to cause extensive degradation of the molecule (6), with the formation of an enzymically active intermediate which can, under controlled conditions, be isolated in almost quantitative yield (7). Pepsin will also cause extensive degradation. Either active (8) or inactive (9) intermediates may be formed, but only the inactive intermediate has been isolated. Although ribonuclease is hydrolyzed only very slowly by carboxypeptidase (lo), it has been observed that impure carboxypeptidase preparations can cause extensive degradation of ribonuclease without a concomitant loss of enzymic activity (11, 12). The identity of the contaminating activities was not determined. However, since neither trypsin nor chymotrypsin degrades native ribonuclease, the results with impure carboxypeptidase suggest the existence of a third pancreatic protease which can degrade native ribonuclease. Elastase has only recently come to be recognized as a distinct member of the family of pancreatic proteolytic enzymes (13). Its specificity has been found to be very broad and different from that of either trypsin or chymotrypsin in that elastase exhibits a relative preference for peptide bonds involving the carboxyl group of aliphatic amino acids (14). Relatively little attention has been paid to the action of elastase on native globular proteins, although its unusual specificity lends interest to such studies. The results of the investigation to be reported in this paper show that elastase is capable of extensively degrading native ribonuclease. The course of the proteolysis offers some unusual features, notably an initial delay, and the accumulation of active intermediates.