title: Deletion of the glycosyltransferase bgsB of Enterococcus faecalis leads to a complete loss of glycolipids from the cell membrane and to impaired biofilm formation.
abstract: BACKGROUND
  Deletion of the glycosyltransferase bgsA in Enterococcus faecalis leads to loss of diglucosyldiacylglycerol from the cell membrane and accumulation of its precursor monoglucosyldiacylglycerol, associated with impaired biofilm formation and reduced virulence in vivo. Here we analyzed the function of a putative glucosyltransferase EF2890 designated biofilm-associated glycolipid synthesis B (bgsB) immediately downstream of bgsA.
  
  
  RESULTS
  A deletion mutant was constructed by targeted mutagenesis in E. faecalis strain 12030. Analysis of cell membrane extracts revealed a complete loss of glycolipids from the cell membrane. Cell walls of 12030ΔbgsB contained approximately fourfold more LTA, and 1H-nuclear magnetic resonance (NMR) spectroscopy suggested that the higher content of cellular LTA was due to increased length of the glycerol-phosphate polymer of LTA. 12030ΔbgsB was not altered in growth, cell morphology, or autolysis. However, attachment to Caco-2 cells was reduced to 50% of wild-type levels, and biofilm formation on polystyrene was highly impaired. Despite normal resistance to cationic antimicrobial peptides, complement and antibody-mediated opsonophagocytic killing in vitro, 12030ΔbgsB was cleared more rapidly from the bloodstream of mice than wild-type bacteria. Overall, the phenotype resembles the respective deletion mutant in the bgsA gene. Our findings suggest that loss of diglucosyldiacylglycerol or the altered structure of LTA in both mutants account for phenotypic changes observed.
  
  
  CONCLUSIONS
  In summary, BgsB is a glucosyltransferase that synthesizes monoglucosyldiacylglycerol. Its inactivation profoundly affects cell membrane composition and has secondary effects on LTA biosynthesis. Both cell-membrane amphiphiles are critical for biofilm formation and virulence of E. faecalis.
authors.affiliation: University Medical Center Freiburg, Germany. christian.theilacker@uniklinik-freiburg.de
authors.forename: Christian
authors.initials: C
authors.lastname: Theilacker
authors.forename: Irina
authors.initials: I
authors.lastname: Sava
authors.forename: Patricia
authors.initials: P
authors.lastname: Sanchez-Carballo
authors.forename: Yinyin
authors.initials: Y
authors.lastname: Bao
authors.forename: Andrea
authors.initials: A
authors.lastname: Kropec
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Grohmann
authors.forename: Otto
authors.initials: O
authors.lastname: Holst
authors.forename: Johannes
authors.initials: J
authors.lastname: Huebner
mesh_terms: D000818:Animals; D016470:Bacteremia / Q000382:microbiology / Q000473:pathology; D001422:Bacterial Adhesion; D058491:Bacterial Load; D018441:Biofilms / Q000254:growth & development*; D018938:Caco-2 Cells; D002462:Cell Membrane / Q000737:chemistry*; D002473:Cell Wall / Q000737:chemistry; D013293:Enterococcus faecalis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D004847:Epithelial Cells / Q000382:microbiology; D005260:Female; D017353:Gene Deletion; D006017:Glycolipids / Q000378:metabolism*; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D016908:Gram-Positive Bacterial Infections / Q000382:microbiology / Q000473:pathology; D006801:Humans; D008070:Lipopolysaccharides / Q000032:analysis; D051379:Mice; D008807:Mice, Inbred BALB C; D013682:Teichoic Acids / Q000032:analysis; D014774:Virulence
chemical_list: D006017:Glycolipids; D008070:Lipopolysaccharides; D013682:Teichoic Acids; C009900:lipoteichoic acid; D016695:Glycosyltransferases