The Primary Structure of Porcine Pancreatic Ribonuclease

Porcine pancreatic ribonuclease contains 8 half-cystine residues located at the same positions in the 124-amino acid residue sequence as in the bovine enzyme. To determine the disposition of the disulfide bonds, the enzyme was subjected to a preliminary proteolysis with pepsin in 5% formic acid and subsequently to the simultaneous action of trypsin and chymotrypsin at pH 6.5. As judged by gel filtration over Sephadex G-25, only negligible quantities of cystine were present in the glycopeptide fraction. Three major cystine-containing fractions obtained by gel filtration were each subjected to fractionation and analysis by zone electrophoresis on paper. They contained five principal cystine peptides. The amino acid compositions of the cysteic acid peptides derived by oxidation of these cystine peptides, together with appropriate sequence analyses, showed that disulfide bonds link half-cystines I–VI, II–VII, III–VIII, and IV–V in the parent molecule.