Structural and Biochemical Characterization of Rm3, a SubClass B3 Metallo-β-Lactamase Identified from a Functional Metagenomic Study
Antimicrobial Agents and Chemotherapy
American Society for Microbiology
Reason for embargo
β-Lactamase production increasingly threatens the effectiveness of β-lactams, which remain a mainstay of antimicrobial chemotherapy. New activities emerge both through mutation of previously known β-lactamases and mobilization from environmental reservoirs. The spread of metallo-β-lactamases (MBLs) represents a particular challenge through their typically broad spectrum activities, encompassing carbapenems in addition to other β-lactam classes. Increasingly, genomic and metagenomic studies reveal distribution of putative MBLs in the environment, but in most cases their activity against clinically relevant β-lactams, and hence the extent to which they can be considered a resistance reservoir, remains uncharacterized. Here we characterize the product of one such gene, blaRm3, identified through functional metagenomic sampling of an environment with high biocide exposure. blaRm3 encodes a subclass B3 MBL that, when expressed in recombinant E. coli, is exported to the bacterial periplasm and hydrolyzes clinically used penicillins, cephalosporins, and carbapenems with an efficiency limited by high KM values. An Rm3 crystal structure reveals the MBL superfamily αβ/βα fold, which more closely resembles mobilized B3 MBLs (AIM-1, SMB-1) than other chromosomal enzymes (L1 or FEZ-1). A binuclear zinc site sits in a deep channel that is in part defined by a relatively extended N-terminus. Structural comparisons suggest that the steric constraints imposed by the N-terminus may limit β-lactam affinity. Sequence comparisons identify Rm3-like MBLs in numerous other environmental samples and species. Our data suggest that Rm3 like enzymes represent a distinct group of B3 MBLs with a wide distribution and can be considered as an environmental reservoir of β-lactam resistance.
R.S., J.S., J.B. and C.J.S. are supported by the U.K. Medical Research Council (U.K.-Canada Team Grant G1100135) and P.H. and J.S. by the National Institute of Allergy and Infectious Diseases of the U.S. National Institutes of Health under award number R01AI100560. LZ, WHG and EMHW were supported by NERC NE/E004482/1 and a European Regional Development Fund, grant no. 500020. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
This is the author accepted manuscript. The final version is available from the American Society for Microbiology via the DOI in this record.
Published online 18 July 2016