IMP-type metallo-β-lactamases (MBLs) are exogenous zinc metalloenzymes that hydrolyze a broad range of β-lactams, including carbapenems. Here we report the crystal structure of IMP-18, an MBL cloned from Pseudomonas aeruginosa, at 2.0-Å resolution. The overall structure of IMP-18 resembles that of IMP-1, with an αβ/βα "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 kcat/Km value increased by an order of magnitude. The results indicate that this is a key residue that defines the kinetic properties of IMP-type β-lactamases.
Submitter: Paulie Dang
Submission Date: March 28, 2020, 5:45 p.m.
Explanation of amino acid numbering: Residues K26N, T32P, and I51F that were mutated in this study are called residues K44N, T50P, and I69F respectively in the paper.
|Number of data points||240|
|Proteins||IMP-18 Crystal Structure|
|Assays/Quantities/Protocols||Experimental Assay: kcat ; Experimental Assay: Km ; Experimental Assay: kcat/Km ; Derived Quantity: SD of kcat/Km ; Derived Quantity: SD of Km ; Derived Quantity: SD of kcat|
|Libraries||Kinetic Assay Data|