In staphylococci and Bacillus,

a single processive glucosyltransferase YpfP adds two glucose residues to DAG to synthesize DGlcDAG [12, 16, 17]. Depending on the bacterial species and strain background, the deletion of this selleck products enzyme may result in an increased LTA content and turnover [16], or loss of LTA from the cell membrane, associated with a reduced rate of selleck autolysis and impaired biofilm formation [12]. In listeria, streptococci, and enterococci, genome analysis revealed two putative glycosyltransferases involved in the biosynthetic pathway of glycolipids [7, 14, 15, 18]. Homologues of a (1→2) glucosyltransferase have been investigated in listeria (LafA), group B streptococci (IagA), and E. faecalis (BgsA) [5, 15, 18]. In group B streptococci, deletion of iagA results in the absence of capsule expression, reduced retention of LTA on the bacterial cell surface, and increased release of LTA into the culture medium [18]. Inactivation of lafA in L. monocytogenes strongly depletes LTA from both the cell wall and the culture medium [18]. In contrast to these findings, deletion of bgsA in E. faecalis results in an increased concentration of LTA in the bacterial cell envelope, most likely related to the longer glycerol-phosphate polymer. The different makeup of glycolipids Dinaciclib mouse and LTA in this mutant

strongly impaired biofilm-formation and affected virulence in vivo [5]. In the current study, we constructed a deletion mutant by targeted mutagenesis of the putative glycosyltransferase bgsB located immediately downstream of bgsA. After inactivation of bgsB in E. faecalis 12030, no glycolipids or glycolipid-derivatives were recovered from the cell envelope of the 12030ΔbgsB mutant, indicating that BgsB is a 1,2-diacylglycerol 3-glucosyltransferase. BgsA cannot take the place of BgsB, which suggests that 4��8C BgsA has higher substrate specificity than YpfP in S. aureus and B. subtilis [13, 17]. The putative function assigned to BgsA and BgsB by this work is in agreement with data obtained for their homologues

LafA and LafB in L. monocytogenes [15]. Although the lipid anchor of LTA from 12030ΔbgsB was not characterized chemically, indirect evidence suggests that DAG instead of DGlcDAG anchors LTA to the cell membrane in this mutant. LTA extracted from 12030ΔbgsB migrated more slowly than wild-type LTA in SDS PAGE, a feature that has been described for homologous LTA molecules substituted with DAG instead of DGlcDAG in S. aureus and L. monocytogenes [13, 15]. In staphylococci and listeria it has been also demonstrated that, in the absence of glycolipids, the enzyme that transfers glycerolphosphate residues to the glycolipid anchor (LtaS) can utilize DAG as glycerolphosphate acceptor for the synthesis of the LTA backbone [13, 15]. Deletion mutants of the glucosyltransferases bgsB and bgsA enabled us to study the individual roles of the two major glycolipids MGlcDAG and DGlcDAG in the physiology and virulence of E. faecalis.