title: Assay platform for clinically relevant metallo-β-lactamases.
abstract: Metallo-β-lactamases (MBLs) are a growing threat to the use of almost all clinically used β-lactam antibiotics. The identification of broad-spectrum MBL inhibitors is hampered by the lack of a suitable screening platform, consisting of appropriate substrates and a set of clinically relevant MBLs. We report procedures for the preparation of a set of clinically relevant metallo-β-lactamases (i.e., NDM-1 (New Delhi MBL), IMP-1 (Imipenemase), SPM-1 (São Paulo MBL), and VIM-2 (Verona integron-encoded MBL)) and the identification of suitable fluorogenic substrates (umbelliferone-derived cephalosporins). The fluorogenic substrates were compared to chromogenic substrates (CENTA, nitrocefin, and imipenem), showing improved sensitivity and kinetic parameters. The efficiency of the fluorogenic substrates was exemplified by inhibitor screening, identifying 4-chloroisoquinolinols as potential pan MBL inhibitors.
authors.affiliation: Chemistry Research Laboratory, University of Oxford , 12 Mansfield Road, Oxford OX1 3TA, United Kingdom.
authors.forename: Sander S
authors.initials: SS
authors.lastname: van Berkel
authors.forename: Jürgen
authors.initials: J
authors.lastname: Brem
authors.forename: Anna M
authors.initials: AM
authors.lastname: Rydzik
authors.forename: Ramya
authors.initials: R
authors.lastname: Salimraj
authors.forename: Ricky
authors.initials: R
authors.lastname: Cain
authors.forename: Anil
authors.initials: A
authors.lastname: Verma
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Owens
authors.forename: Colin W G
authors.initials: CW
authors.lastname: Fishwick
authors.forename: James
authors.initials: J
authors.lastname: Spencer
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Schofield
mesh_terms: D004791:Enzyme Inhibitors / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008670:Metals / Q000737:chemistry*; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases / Q000032:analysis* / Q000737:chemistry
chemical_list: D004791:Enzyme Inhibitors; D008670:Metals; D065093:beta-Lactamase Inhibitors; D001618:beta-Lactamases
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