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DNA Gyrase from the Albicidin Producer Xanthomonas albilineans Has Multiple-Antibiotic-Resistance and Unusual Enzymatic Properties

Saeed M. Hashimi,¹ Guozhong Huang,^1, Anthony Maxwell,² and Robert G. Birch^1*

Botany Department—SIB, The University of Queensland, Brisbane 4072, Australia,¹ Department of Biological Chemistry, John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, United Kingdom²

Received 2 December 2007/ Returned for modification 4 January 2008/ Accepted 29 January 2008

The sugarcane pathogen Xanthomonas albilineans produces a family of antibiotics and phytotoxins termed albicidins, which inhibit plant and bacterial DNA gyrase supercoiling activity, with a 50% inhibitory concentration (50 nM) comparable to those of coumarins and quinolones. Here we show that X. albilineans has an unusual, antibiotic-resistant DNA gyrase. The X. albilineans gyrA and gyrB genes are not clustered with previously described albicidin biosynthesis and self-protection genes. The GyrA and GyrB products differ from Escherichia coli homologues through several insertions and through changes in several amino acid residues implicated in quinolone and coumarin resistance. Reconstituted X. albilineans DNA gyrase showed 20- to 25-fold-higher resistance than E. coli DNA gyrase to albicidin and ciprofloxacin and 8-fold-higher resistance to novobiocin in the supercoiling assay. The X. albilineans DNA gyrase is unusual in showing a high degree of distributive supercoiling and little DNA relaxation activity. X. albilineans GyrA (XaA) forms a functional gyrase heterotetramer with E. coli GyrB (EcB) and can account for albicidin and quinolone resistance and low levels of relaxation activity. XaB probably contributes to both coumarin resistance and the distributive supercoiling pattern. Although XaB shows fewer apparent changes relative to EcB, the EcA·XaB hybrid relaxed DNA in the presence or absence of ATP and was unable to supercoil. A fuller understanding of structural differences between albicidin-sensitive and -resistant gyrases may provide new clues into features of the enzyme amenable to interference by novel antibiotics.

Published ahead of print on 11 February 2008.

Present address: Department of Plant Pathology, The University of Georgia, Athens, GA 30602-7274.