Systematic Analysis of Metallo-β-Lactamases Using an Automated Database

Abstract

b Metallo-␤-lactamases (MBLs) are enzymes that hydrolyze ␤-lactam antibiotics, resulting in bacterial resistance to these drugs. These proteins have caused concerns due to their facile transference, broad substrate spectra, and the absence of clinically useful inhibitors. To facilitate the classification, nomenclature, and analysis of MBLs, an automated database system was developed, the Metallo-␤-Lactamase Engineering Database (MBLED) (http://www.mbled.uni-stuttgart.de). It contains information on MBLs retrieved from the NCBI peptide database while strictly following the nomenclature by Jacoby and Bush (http://www.lahey.org /Studies/) and the generally accepted class B ␤-lactamase (BBL) standard numbering scheme for MBLs. The database comprises 597 MBL protein sequences and enables systematic analyses of these sequences. A systematic analysis employing the database resulted in the generation of mutation profiles of assigned IMP-and VIM-type MBLs, the identification of five MBL protein entries from the NCBI peptide database that were inconsistent with the Jacoby and Bush nomenclature, and the identification of 15 new IMP candidates and 9 new VIM candidates. Furthermore, the database was used to identify residues with high mutation frequencies and variability (mutation hot spots) that were unexpectedly distant from the active site located in the ␤␤ sandwich: positions 208 and 266 in the IMP family and positions 215 and 258 in the VIM family. We expect that the MBLED will be a valuable tool for systematically cataloguing and analyzing the increasing number of MBLs being reported.

Carbapenemase-Producing Enterobacteriaceae Recovered from the Environment of a Swine Farrow-to-Finish Operation in the United States

“…A third P. mirabilis isolate harboring bla IMP-27 was recovered from a patient in the U.S. upper plains region in 2015 (48). bla IMP-27 differs from the first reported IMP-1 by 50 amino acid substitutions (25) and from its closest relative, IMP-8, by 31 amino acid substitutions (23).…”

Detection of IMP Metallo-β-Lactamase in Carbapenem-Nonsusceptible Enterobacteriaceae and Non-Glucose-Fermenting Gram-Negative Rods by Immunochromatography Assay

“…Two monoclonal antibodies designated 4C9-C/F6 and 4E7-C/F6 are used in this IC assay system (14). Since 4C9-C/F6 and 4E7-C/F6 recognize the highly conserved amino acids 124 to 130 (the H2 region) and 134 to 140 (the S6 region) in IMP (14,24), and since the H2 and S6 regions of the new IMP types, IMP-40, -41, and -42, are well conserved, the IC assay detects such IMP types. Thus, the IC assay might also be used for detecting emerging types of IMP that retain the original H2 and S6 regions.…”

“…Amplicons obtained from each PCR were sequenced using M13F and M13R primers, the BigDye Terminator v3.1 Cycle Sequencing Kit (ABI, Carlsbad, CA), and an ABI sequence analyzer 3730XL (ABI). Each type of IMP was determined by a BLAST search and data based on all the bla IMP sequencing described previously (http://www .lahey.org/studies/other.asp#Table 1) (24).…”

New β-Lactamase Inhibitors: a Therapeutic Renaissance in an MDR World

“…(79)(80)(81). As these enzymes have "grown up" during this technological age, multiple computer-based approaches, such as computer modeling of target compounds, integrating protein shape and surface charge profiles into in silico screening, and compiling crystal structures in automated databases for future studies, have been described for drug discovery (82)(83)(84).…”