Identification of vancomycin resistance protein VanA as a D-alanine:D-alanine ligase of altered substrate specificity.

High-level glycopeptide resistance in Enterococcus faecium BM4147 is mediated by a 38-kDa protein VanA, whose amino acid sequence is related to Gram-negative D-alanine:D-alanine (D-Ala-D-Ala) ligases [Dutka-Malen, S., Molinas, C., Arthur, M., & Courvalin, P. (1990) Mol. Gen. Genet. 224, 364-372]. We report purification of VanA and demonstrate that it has D-Ala-D-Ala ligase activity but has substantially modified substrate specificity, compared with Gram-negative D-Ala-D-Ala ligases. VanA preferentially condenses D-Ala with D-Met or D-Phe, raising the possibility that its cellular role is to synthesize a modified cell-wall component, which is subsequently not recognized by vancomycin.