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Nucleic Acids Res. 2013 Apr;41(8):4628-39. doi: 10.1093/nar/gkt124. Epub 2013 Mar 4.

Topoisomerase IV-quinolone interactions are mediated through a water-metal ion bridge: mechanistic basis of quinolone resistance.

Author information

1
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA.

Abstract

Although quinolones are the most commonly prescribed antibacterials, their use is threatened by an increasing prevalence of resistance. The most common causes of quinolone resistance are mutations of a specific serine or acidic residue in the A subunit of gyrase or topoisomerase IV. These amino acids are proposed to serve as a critical enzyme-quinolone interaction site by anchoring a water-metal ion bridge that coordinates drug binding. To probe the role of the proposed water-metal ion bridge, we characterized wild-type, GrlA(E85K), GrlA(S81F/E85K), GrlA(E85A), GrlA(S81F/E85A) and GrlA(S81F) Bacillus anthracis topoisomerase IV, their sensitivity to quinolones and related drugs and their use of metal ions. Mutations increased the Mg(2+) concentration required to produce maximal quinolone-induced DNA cleavage and restricted the divalent metal ions that could support quinolone activity. Individual mutation of Ser81 or Glu85 partially disrupted bridge function, whereas simultaneous mutation of both residues abrogated protein-quinolone interactions. Results provide functional evidence for the existence of the water-metal ion bridge, confirm that the serine and glutamic acid residues anchor the bridge, demonstrate that the bridge is the primary conduit for interactions between clinically relevant quinolones and topoisomerase IV and provide a likely mechanism for the most common causes of quinolone resistance.

PMID:
23460203
PMCID:
PMC3632122
DOI:
10.1093/nar/gkt124
[Indexed for MEDLINE]
Free PMC Article

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