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Antimicrobial Agents and Chemotherapy, October 2002, p. 3125-3132, Vol. 46, No. 10
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.10.3125-3132.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Biochemical and Genetic Characterization of the vanC-2 Vancomycin Resistance Gene Cluster of Enterococcus casseliflavus ATCC 25788

Ireena Dutta and Peter E. Reynolds^*

Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom CB2 1QW

Received 27 July 2001/ Returned for modification 21 September 2001/ Accepted 17 May 2002

The vanC-2 cluster of Enterococcus casseliflavus ATCC 25788 consisted of five genes (vanC-2, vanXY_C-2, vanT_C-2, vanR_C-2, and vanS_C-2) and shared the same organization as the vanC cluster of E. gallinarum BM4174. The proteins encoded by these genes displayed a high degree of amino acid identity to the proteins encoded within the vanC gene cluster. The putative D,D-dipeptidase-D,D-carboxypeptidase, VanXY_C-2, exhibited 81% amino acid identity to VanXY_C, and VanT_C-2 displayed 65% amino acid identity to the serine racemase, VanT. VanR_C-2 and VanS_C-2 displayed high degrees of identity to VanR_C and VanS_C, respectively, and contained the conserved residues identified as important to their function as a response regulator and histidine kinase, respectively. Resistance to vancomycin was expressed inducibly in E. casseliflavus ATCC 25788 and required an extended period of induction. Analysis of peptidoglycan precursors revealed that UDP-N-acetylmuramyl-L-Ala--D-Glu-L-Lys-D-Ala-D-Ser could not be detected until several hours after the addition of vancomycin, and its appearance coincided with the resumption of growth. The introduction of additional copies of the vanT_C-2 gene, encoding a putative serine racemase, and the presence of supplementary D-serine in the growth medium both significantly reduced the period before growth resumed after addition of vancomycin. This suggested that the availability of D-serine plays an important role in the induction process.

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* Corresponding author. Mailing address: Department of Biochemistry, University of Cambridge, Tennis Court Rd., Cambridge, United Kingdom CB2 1QW. Phone: (44) 1223 740494. Fax: (44) 1223 333345. E-mail: per{at}mole.bio.cam.ac.uk.

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Antimicrobial Agents and Chemotherapy, October 2002, p. 3125-3132, Vol. 46, No. 10
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.10.3125-3132.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Reynolds, P. E., Courvalin, P. (2005). Vancomycin Resistance in Enterococci Due to Synthesis of Precursors Terminating in D-Alanyl-D-Serine. Antimicrob. Agents Chemother. 49: 21-25 [Full Text]  
• Arias, C. A., Pena, J., Panesso, D., Reynolds, P. (2003). Role of the transmembrane domain of the VanT serine racemase in resistance to vancomycin in Enterococcus gallinarum BM4174. J Antimicrob Chemother 51: 557-564 [Abstract] [Full Text]  
• Dutta, I., Reynolds, P. E. (2003). The vanC-3 vancomycin resistance gene cluster of Enterococcus flavescens CCM 439. J Antimicrob Chemother 51: 703-706 [Abstract] [Full Text]