Structural and Mutagenic Analysis of Metallo-β-Lactamase IMP-18

IMP-type metallo-beta-lactamases (MBLs) are exogenous zinc metalloenzymes that hydrolyze a broad range of beta-lactams, including carbapenems. Here we report the crystal structure of IMP-18, an MBL cloned from Pseudomonas aeruginosa, at 2.0-angstrom resolution. The overall structure of IMP-18 resembles that of IMP-1, with an alpha beta/beta alpha folded sandwich configuration, but the loop that covers the active site has a distinct conformation. The relationship between IMP-18' s loop conformation and its kinetic properties was investigated by replacing the amino acid residues that can affect the loop conformation (Lys44, Thr50, and Ile69) in IMP-18 with those occupying the corresponding positions in the well-described enzyme IMP-1. The replacement of Thr50 with Pro considerably modified IMP-18' s kinetic properties, specifically those pertaining to meropenem, with the k(cat)/K-m value increased by an order of magnitude. The results indicate that this is a key residue that defines the kinetic properties of IMP-type beta-lactamases.