Crystal structure of Pseudomonas aeruginosa SPM-1 provides insights into variable zinc affinity of metallo-β-lactamases

Tanya A. Murphy, Lucy E. Catto, Stephen E. Halford, Andrea T. Hadfield, Wladek Minor, Timothy R. Walsh, James Spencer

Research output: Contribution to journalArticlepeer-review

74 Scopus citations

Abstract

Metallo-β-lactamases (mβls) confer broad-spectrum resistance to β-lactam antibiotics upon host bacteria and escape the action of existing β-lactamase inhibitors. SPM-1 is a recently discovered mβl that is distinguished from related enzymes by possession of a substantial central insertion and by sequence variation at positions that maintain active site structure. Biochemical data show SPM-1 to contain two Zn2+ sites of differing affinities, a phenomenon that is well documented amongst mβls but for which a structural explanation has proved elusive. Here, we report the crystal structure of SPM-1 to 1.9 Å resolution. The structure reveals SPM-1 to lack a mobile loop implicated in substrate binding by related mβls and to accommodate the central insertion in an extended helical interdomain region. Deleting this had marginal effect upon binding and hydrolysis of a range of β-lactams. These data suggest that the interactions of SPM-1 with substrates differ from those employed by other mβls. SPM-1 as crystallised contains a single Zn2+. Both the active site hydrogen-bonding network and main-chain geometry at Asp120, a key component of the binding site for the second zinc ion, differ significantly from previous mβl structures. We propose that variable interactions made by the Asp120 carbonyl group modulate affinity for a second Zn2+ equivalent in mβls of the B1 subfamily. We further predict that SPM-1 possesses the capacity to evolve variants of enhanced catalytic activity by point mutations altering geometry and hydrogen bonding in the vicinity of the second Zn2+ site.

Original languageEnglish
Pages (from-to)890-903
Number of pages14
JournalJournal of Molecular Biology
Volume357
Issue number3
DOIs
StatePublished - 31 Mar 2006
Externally publishedYes

Keywords

  • Antibiotic resistance
  • Dinuclear zinc
  • Hydrolase
  • Metallo-β-lactamase
  • Structure

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