title: Targeting metallo-carbapenemases via modulation of electronic properties of cephalosporins.
abstract: The global proliferation of metallo-carbapenemase-producing Enterobacteriaceae has created an unmet need for inhibitors of these enzymes. The rational design of metallo-carbapenemase inhibitors requires detailed knowledge of their catalytic mechanisms. Nine cephalosporins, structurally identical except for the systematic substitution of electron-donating and withdrawing groups in the para position of the styrylbenzene ring, were synthesized and utilized to probe the catalytic mechanism of New Delhi metallo-β-lactamase (NDM-1). Under steady-state conditions, K(m) values were all in the micromolar range (1.5-8.1 μM), whereas k(cat) values varied widely (17-220 s(-1)). There were large solvent deuterium isotope effects for all substrates under saturating conditions, suggesting a proton transfer is involved in the rate-limiting step. Pre-steady-state UV-visible scans demonstrated the formation of short-lived intermediates for all compounds. Hammett plots yielded reaction constants (ρ) of -0.34 ± 0.02 and -1.15 ± 0.08 for intermediate formation and breakdown, respectively. Temperature-dependence experiments yielded ΔG(‡) values that were consistent with the Hammett results. These results establish the commonality of the formation of an azanide intermediate in the NDM-1-catalysed hydrolysis of a range cephalosporins with differing electronic properties. This intermediate is a promising target for judiciously designed β-lactam antibiotics that are poor NDM-1 substrates and inhibitors with enhanced active-site residence times.
authors.affiliation: *Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056, U.S.A.
authors.forename: Hao
authors.initials: H
authors.lastname: Yang
authors.affiliation: *Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056, U.S.A.
authors.forename: Heather
authors.initials: H
authors.lastname: Young
authors.affiliation: †Sopharmia Inc, Christopher S "Kit" Bond Science and Technology Incubator, Missouri Western State University, 4221 Mitchell Ave, Saint Joseph, MO 64507, U.S.A.
authors.forename: Sophia
authors.initials: S
authors.lastname: Yu
authors.forename: Larry
authors.initials: L
authors.lastname: Sutton
authors.affiliation: *Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056, U.S.A.
authors.forename: Michael W
authors.initials: MW
authors.lastname: Crowder
mesh_terms: D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D002511:Cephalosporins / Q000737:chemistry* / Q000494:pharmacology; D004755:Enterobacteriaceae / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D018440:beta-Lactam Resistance / Q000187:drug effects; D001618:beta-Lactamases / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D002511:Cephalosporins; D004791:Enzyme Inhibitors; D001618:beta-Lactamases; C552864:beta-lactamase NDM-1; C063912:carbapenemase