Dataset Information

VanC cluster of vancomycin-resistant Enterococcus gallinarum BM4174.

ABSTRACT: Glycopeptide-resistant enterococci of the VanC type synthesize UDP-muramyl-pentapeptide[D-Ser] for cell wall assembly and prevent synthesis of peptidoglycan precursors ending in D-Ala. The vanC cluster of Enterococcus gallinarum BM4174 consists of five genes: vanC-1, vanXY(C), vanT, vanR(C), and vanS(C). Three genes are sufficient for resistance: vanC-1 encodes a ligase that synthesizes the dipeptide D-Ala-D-Ser for addition to UDP-MurNAc-tripeptide, vanXY(C) encodes a D,D-dipeptidase-carboxypeptidase that hydrolyzes D-Ala-D-Ala and removes D-Ala from UDP-MurNAc-pentapeptide[D-Ala], and vanT encodes a membrane-bound serine racemase that provides D-Ser for the synthetic pathway. The three genes are clustered: the start codons of vanXY(C) and vanT overlap the termination codons of vanC-1 and vanXY(C), respectively. Two genes which encode proteins with homology to the VanS-VanR two-component regulatory system were present downstream from the resistance genes. The predicted amino acid sequence of VanR(C) exhibited 50% identity to VanR and 33% identity to VanR(B). VanS(C) had 40% identity to VanS over a region of 308 amino acids and 24% identity to VanS(B) over a region of 285 amino acids. All residues with important functions in response regulators and histidine kinases were conserved in VanR(C) and VanS(C), respectively. Induction experiments based on the determination of D,D-carboxypeptidase activity in cytoplasmic extracts confirmed that the genes were expressed constitutively. Using a promoter-probing vector, regions upstream from the resistance and regulatory genes were identified that have promoter activity.

SUBMITTER: Arias CA

PROVIDER: S-EPMC89929 | biostudies-literature | 2000 Jun

REPOSITORIES: biostudies-literature

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vanC cluster of vancomycin-resistant Enterococcus gallinarum BM4174.

Arias C A CA Courvalin P P Reynolds P E PE

Antimicrobial agents and chemotherapy 20000601 6

Glycopeptide-resistant enterococci of the VanC type synthesize UDP-muramyl-pentapeptide[D-Ser] for cell wall assembly and prevent synthesis of peptidoglycan precursors ending in D-Ala. The vanC cluster of Enterococcus gallinarum BM4174 consists of five genes: vanC-1, vanXY(C), vanT, vanR(C), and vanS(C). Three genes are sufficient for resistance: vanC-1 encodes a ligase that synthesizes the dipeptide D-Ala-D-Ser for addition to UDP-MurNAc-tripeptide, vanXY(C) encodes a D,D-dipeptidase-carboxypep ...[more]

PMID: 10817725

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Project description:Enterococcus faecium clinical isolates A902 and BM4538, which were resistant to relatively high levels of vancomycin (128 and 64 microg/ml, respectively) and to low levels of teicoplanin (4 microg/ml), and Enterococcus faecalis clinical isolates BM4539 and BM4540, which were resistant to moderate levels of vancomycin (16 microg/ml) and susceptible to teicoplanin (0.25 microg/ml), were studied. They were constitutively resistant by synthesis of peptidoglycan precursors ending with d-alanyl-d-lactate and harbored a chromosomal vanD gene cluster which was not transferable by conjugation to other enterococci. VanX(D) activity, which is not required in the absence of d-Ala-d-Ala, was low in the four strains, although none of the conserved residues was mutated; and the constitutive VanY(D) activity in the membrane fractions was inhibited by penicillin G. The mutations E(13)G in the region of d-alanine:d-alanine ligase (which is implicated in d-Ala1 binding in A902) and S(319)N of the serine involved in ATP binding in BM4538 and a 7-bp insertion at different locations in BM4539 and BM4540 (which led to putative truncated proteins) led to the production of an impaired enzyme and accounted for the lack of d-Ala-d-Ala-containing peptidoglycan precursors. The same 7-bp insertion in vanS(D) of BM4539 and BM4540 and a 1-bp deletion in vanS(D) of A902, which in each case led to a putative truncated and presumably nonfunctional protein, could account for the constitutive resistance. Strain BM4538, with a functional VanS(D), had a G(140)E mutation in VanR(D) that could be responsible for constitutive glycopeptide resistance. This would represent the first example of constitutive van gene expression due to a mutation in the structural gene for a VanR transcriptional activator. Study of these four additional strains that could be distinguished on the basis of their various assortments of mutations confirmed that all VanD-type strains isolated so far have mutations in the ddl housekeeping gene and in the acquired vanS(D) or vanR(D) gene that lead to constitutive resistance to vancomycin.

Dataset Information

VanC cluster of vancomycin-resistant Enterococcus gallinarum BM4174.

Publications

vanC cluster of vancomycin-resistant Enterococcus gallinarum BM4174.

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