The Structure of the Porcine Pancreatic Secretory Trypsin Inhibitor I

Porcine pancreatic secretory trypsin inhibitor I (56 residues) was rendered sensitive to tryptic digestion by reduction and carboxymethylation. Nine tryptic fragments were formed by the action of trypsin. They were separated by taking advantage of the size fractionating and ion exchange properties of Bio Gel P‐2 in 0.01 N pyridine. The major tryptic peptides were purified on SE‐Sephadex C‐25. The amino acid sequence of each peptide was determined by a special technique of Edman degradation using p‐bromophenyl‐isothiocyanate. This reagent facilitated the identification of the extracted p‐bromophenyl‐thiohydantoines by mass spectrometry. The characteristic fragment and molecular ions are emphasized by a significant double peak of a mass difference Δm/e= 2, which is caused by the isotope ratio of the p‐bromosubstituent. In general this technique permitted up to thirteen consecutive steps of degradation and the unequivocal positioning of the amides. The complete amino acid sequence of the polypeptide trypsin inhibitor was determined.