An acidic glutaryl-7-aminocephalosporanic acid acylase from Pseudomonas nitroreducens.

A glutaryl-7-aminocephalosporanic acid (GL-7-ACA) acylase was purified 58-fold from Pseudomonas nitroreducens in a two-step procedure involving osmotic shock and carboxymethyl-Sepharose chromatography with a yield of 26%. The molecular mass of the native enzyme was 58 kDa. SDS/PAGE revealed that it consisted of two non-identical subunits with molecular masses of 35 and 21 kDa. The isoelectric point of the purified enzyme was 5.3. The enzyme had an optimal pH of 5.5 and an optimal temperature of 43 degrees C. The purified enzyme exhibited not only GL-7-ACA acylase activity but also gamma-glutamyltranspeptidase activity. The Km values of the enzyme for GL-7-ACA and L-gamma-glutamyl p-nitroanilide were 10.41 mM and 5.92 microM respectively.