Disrupting antibiotic resistance propagation by inhibiting the conjugative DNA relaxase

doi:10.1073/pnas.0702760104

Disrupting antibiotic resistance propagation by inhibiting the conjugative DNA relaxase

Departments of *Chemistry,
^†Biochemistry and Biophysics, and
^‡Biology,
^§Curriculum in Genetics and Molecular Biology, and
^¶Program in Molecular Biology and Biotechnology, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599-3290

Edited by Donald R. Helinski, University of California at San Diego, La Jolla, CA, and approved June 11, 2007 (received for review March 23, 2007)

Abstract

Conjugative transfer of plasmid DNA via close cell–cell junctions is the main route by which antibiotic resistance genes spread between bacterial strains. Relaxases are essential for conjugative transfer and act by cleaving DNA strands and forming covalent phosphotyrosine linkages. Based on data indicating that multityrosine relaxase enzymes can accommodate two phosphotyrosine intermediates within their divalent metal-containing active sites, we hypothesized that bisphosphonates would inhibit relaxase activity and conjugative DNA transfer. We identified bisphosphonates that are nanomolar inhibitors of the F plasmid conjugative relaxase in vitro. Furthermore, we used cell-based assays to demonstrate that these compounds are highly effective at preventing DNA transfer and at selectively killing cells harboring conjugative plasmids. Two potent inhibitors, clodronate and etidronate, are already clinically approved to treat bone loss. Thus, the inhibition of conjugative relaxases is a potentially novel antimicrobial approach, one that selectively targets bacteria capable of transferring antibiotic resistance and generating multidrug resistant strains.

Footnotes

^‖To whom correspondence should be addressed. E-mail: redinbo{at}unc.edu
Author contributions: S.A.L., L.M.G., S.W.M., and M.R.R. designed research; S.A.L., L.M.G., and H.R. performed research; S.A.L. analyzed data; and S.A.L. and M.R.R. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
Data deposition: The atomic coordinates for the structures without and with PNP have been deposited in the Protein Data Bank, www.pdb.org (PDB ID codes 2Q7T and 2Q7U, respectively).
This article contains supporting information online at www.pnas.org/cgi/content/full/0702760104/DC1.
Abbreviations:

PCP,

methylenediphosphonic acid;

PCNCP,

iminobis(methylphosphonic acid);

ETIDRO,

etidronic acid;

CLODRO,

clodronic acid;

PBENP,

1,2-bis(dimethoxyphosphoryl)benzene;

PNP,

imidobisphosphate;

CPR,

concomitant plasmid replication.
Freely available online through the PNAS open access option.