Preparation of dipeptidyl aminopeptidase IV for polypeptide sequencing

Dipeptidyl aminopeptidase IV is a dipeptidylpeptide hydrolase (EC 3.4.14) which hydrolyzes bond at the carboxyl group of proline releasing X-Pro dipeptides from the amino-terminus of polypeptides. The enzyme was purified 440-fold in 37% yield from swine kidney by ammonium sulfate fractionation, DEAE-cellulose chromatography, gel filtration and affinity chromatography with dipeptide-substituted Sepharose 4B. The enzyme released X-Pro from all X-Pro-beta-naphthylamides and polypeptides tested. The released dipeptides were not further degraded, and were readily identified in digests. The enzyme is suitable for use in the dipeptidyl aminopeptidase method for sequence analysis of polypeptides.