The Purification and Properties of Penicillin β-Lactamases Mediated by Transmissible R Factors in Escherichia coli

Abstract

1. Two types of penicillin β-lactamases [EC 3.5.2.6], penicillinase,₈₈ and penicillinase₁₄, were isolated from E. coli harboring transmissible R factors, R_{GN235 and RGN14 respectively, and purified mainly by means of column chromatography. Penicillinase238 belonging to type II-penicillinases was purified about 158 fold, and penicillinase14, which is one of type I-penicillinases was purified about 270 fold. 2. Isoelectric point of penicillinase238 determined by agar gel electrophoresis was 8.3, and its s20, w was 2.66S. The optimal pH was around 7.6 and the optimal temperature was 30°C for the hydrolysis of benzylpenicillin. 3. With respect to penicillinase14 the isoelectric point was 5.1 and the value of s20, w was 1.43S in the synthetic boundary cell. The maximal enzyme activity to benzylpenicillin was observed at 45°C and at pH 6.5. 4. The difference between the two enzymes was also revealed by the behaviors to anions like chloride and the kinetic aspects (Km and Vmax).}