The Elastase‐Like Enzymes from Streptomyces griseus (Pronase): Isolation and Partial Characterization

Three elastase‐like enzymes, designated Enzyme I, II and III, were purified from a commercial preparation of Streptomyces griseus (pronase) by successive chromatography on DEAE‐cellulose at pH 9.5 and CM‐cellulose at pH 5.5. Electrophoresis on cellulose acetate at pH 6.9, on polyacrylamide gels at pH 4.5 and 9.5 and exclusion chromatography on Sephadex G‐75 indicated that all three fractions were essentially homogenous. The amino acid composition of the three enzymes was determined and their molecular weights were estimated by gel‐chromatographic methods. All three enzymes possess strong esterolytic activity when assayed on a specific elastase substrate N‐acetyl‐L‐alanyl‐L‐alanyl‐L‐alanine methyl ester (AcAla₃OMe) and are active, to a smaller extent, on N‐acetyl‐L‐tyrosine ethyl ester. Their activity on α‐N‐benzoyl‐L‐arginine ethyl ester was negligible. All the enzymes have strong proteolytic activity at slight alkaline pH and Enzyme II and III possess also an elastolytic activity on congo red elastin. All three enzymes are inhibited by diisopropylfluorophosphate. Enzymes II and III were slowly inhibited by N‐tosyl‐L‐phenylalanine chloromethane. Enzyme I was inhibited by the esterolysis products of AcAla₃OMe, thus indicating that the same active site controls both esterolytic activities.