Abstract
Penicillin acylase (EC 3. 5. 1. 11) of Kluyvera citrophila KY7844 was purified approximately 120-fold by DEAE-cellulose chromatography, hydroxyapatite chromatography and isoelectrofocusing fractionation. The purified enzyme, with an approximate molecular weight of 63, 000, appeared to be homogeneous in disc electrophoretic analysis, and showed isoelectric point (Ip) 8.12 and 13.0 units/mg of specific activity for cephalexin hydrolysis. The Michaelis constant (Km) for cephalexin and for 7-[1-(1H)-tetrazolylacetamido]-desacetoxycephalosporanic acid ((1H) T-7ADCA) was 1.4mm and 3.6mm, respectively. This enzyme was capable of producing (1H) T-7ADCA in 80% yield from 1-(1H)-tetrazolylacetate methylester and 7-aminodesaceto-xycephalosporanic acid.
