Purification and characterization of Prevotella intermedia β-lactamase

Abstract

I studied the effects of β-lactamase inhibitor (tazobactam, clavulanic acid and sulbactam) against β-lactamase, and investigated characterization of β-lactamase to analyze the resistance mechanism of β-lactams in Prevotella intermedia. The activity of β-lactamase, which originated in P. intermedia MA-l and its variants (v1-v11), was blocked by three inhibitors both in vivo and in vitro. β-Lactamase was purified by ion exchange column chromatography (DEAE DE-52) and gel filtration (Sephacryl S-100HR). β-Lactamase at a second peak was eluted from Sephacryl. This enzyme also showed only one peak at retention time 6'18" by HPLC (TSK gel, 2000SW column). The purified enzyme had a molecular weight of 42kDa, optimum pH of 6.0, and an isoelectric point of 8.0. Its enzyme activity was blocked by Hg^<2+>. These results suggest that P. intermedia β-lactamase is a type C cephalosporinase.