Studies on Ribonuclease S

Specific regions of the polypeptide sequence of pancreatic ribonuclease have been altered by chemical modification and by limited proteolytic digestion in attempts to implicate specific covalent portions of the molecule in the structure and stabilization of the active center. Digestion of the native molecule with trypsin at elevated temperatures has been shown to remove portions of the chain with only partial inactivation (1). Conversely, it has been possible to cause total inactivation of the enzyme through removal of the COOH-terminal tetrapeptide sequence, Asp-Ala-Ser-Val, by pepsin (2) or the 20-amino acid fragment at the NHt-terminal end of the molecule by treatment with subtilisin (3). The present studies were designed to analyze the specific roles of individual amino acid residues within the essential NH,and COOH-terminal regions of ribonuclease. In contrast to native RNase A (4), RNase S and its separated components, RNase S-protein and RNase S-peptide, are susceptible to extensive digestion by carboxypeptidase. The experiments described below show that changes in the conformation and the enzymic function of the RNase S derivatives produced can be related to the removal of individual amino acid residues.