A Chymotrypsin-like Protease from the Rat Submandibular Gland.

Abstract

A chymotrypsin-like protease was obtained from the rat submandibular gland by precipitation at pH 5.2, ammonium sulphate fractionation, gel filtration on Sephadex G-200 and DEAE-cellulose chromatography. The degree of purity was increased 270 times. By starch gel electrophoresis at pH 6.0 of the final preparation two enzymatically active protein components moving to the anode were separated. The purified enzyme had a molecular weight of 19 300, and a pH optimum at 7.8 with different substrates. It readily hydrolyzed human hemoglobin, N-CBZ-L-phenylalanine p-nitroanilide, N-CBZ-L-tyrosine hydrazide, acetyl-L-tyrosine ethyl ester, acetyl-L-phenylalanine ethyl ester and benzoyl-DL-phenylalanine [beta]-naphythl ester. Arylamides of unsubstituted aromatic amino-acids were hydrolyzed more slowly. Liberation of tyrosine from poly-L-tryrosine and ammonia from glycyl-DL-phenylalanine amide and glycyl-L-tyrosine amide was also demonstrated. The hydrolytic activity of the enzyme was stimulated by Ca2+ and was not affected by a number of modifying agents.