Main content area

A Noncanonical Metal Center Drives the Activity of the Sediminispirochaeta smaragdinae Metallo-β-lactamase SPS-1

Cheng, Zishuo, VanPelt, Jamie, Bergstrom, Alexander, Bethel, Christopher, Katko, Andrew, Miller, Callie, Mason, Kelly, Cumming, Erin, Zhang, Huan, Kimble, Robert L., Fullington, Sarah, Bretz, Stacey Lowery, Nix, Jay C., Bonomo, Robert A., Tierney, David L., Page, Richard C., Crowder, Michael W.
Biochemistry 2018 v.57 no.35 pp. 5218-5229
active sites, beta-lactamase, binding sites, carbapenems, cephalosporins, cobalt, crystal structure, histidine, hydrogen bonding, metal ions, spectral analysis, spectroscopy, zinc
In an effort to evaluate whether a recently reported putative metallo-β-lactamase (MβL) contains a novel MβL active site, SPS-1 from Sediminispirochaeta smaragdinae was overexpressed, purified, and characterized using spectroscopic and crystallographic studies. Metal analyses demonstrate that recombinant SPS-1 binds nearly 2 equiv of Zn(II), and steady-state kinetic studies show that the enzyme hydrolyzes carbapenems and certain cephalosporins but not β-lactam substrates with bulky substituents at the 6/7 position. Spectroscopic studies of Co(II)-substituted SPS-1 suggest a novel metal center in SPS-1, with a reduced level of spin coupling between the metal ions and a novel Zn₁ metal binding site. This site was confirmed with a crystal structure of the enzyme. The structure shows a Zn₂ site that is similar to that in NDM-1 and other subclass B1 MβLs; however, the Zn₁ metal ion is coordinated by two histidine residues and a water molecule, which is held in position by a hydrogen bond network. The Zn₁ metal is displaced nearly 1 Å from the position reported in other MβLs. The structure also shows extended helices above the active site, which create a binding pocket that precludes the binding of substrates with large, bulky substituents at the 6/7 position of β-lactam antibiotics. This study reveals a novel metal binding site in MβLs and suggests that the targeting of metal binding sites in MβLs with inhibitors is now more challenging with the identification of this new MβL.