title: Identification of Poly-N-acetylglucosamine as a Major Polysaccharide Component of the Bacillus subtilis Biofilm Matrix.
abstract: Bacillus subtilis is intensively studied as a model organism for the development of bacterial biofilms or pellicles. A key component is currently undefined exopolysaccharides produced from proteins encoded by genes within the eps locus. Within this locus are four genes, epsHIJK, known to be essential for pellicle formation. We show they encode proteins synthesizing the broadly expressed microbial carbohydrate poly-N-acetylglucosamine (PNAG). PNAG was present in both pellicle and planktonic wild-type B. subtilis cells and in strains with deletions in the epsA-G and -L-O genes but not in strains deleted for epsH-K. Cloning of the B. subtilis epsH-K genes into Escherichia coli with in-frame deletions in the PNAG biosynthetic genes pgaA-D, respectively, restored PNAG production in E. coli. Cloning the entire B. subtilis epsHIJK locus into pga-deleted E. coli, Klebsiella pneumoniae, or alginate-negative Pseudomonas aeruginosa restored or conferred PNAG production. Bioinformatic and structural predictions of the EpsHIJK proteins suggest EpsH and EpsJ are glycosyltransferases (GT) with a GT-A fold; EpsI is a GT with a GT-B fold, and EpsK is an α-helical membrane transporter. B. subtilis, E. coli, and pga-deleted E. coli carrying the epsHIJK genes on a plasmid were all susceptible to opsonic killing by antibodies to PNAG. The immunochemical and genetic data identify the genes and proteins used by B. subtilis to produce PNAG as a significant carbohydrate factor essential for pellicle formation.
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and.
authors.forename: Damien
authors.initials: D
authors.lastname: Roux
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and.
authors.forename: Colette
authors.initials: C
authors.lastname: Cywes-Bentley
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and Harvard School of Dental Medicine, Boston, Massachusetts 02115.
authors.forename: Yi-Fan
authors.initials: YF
authors.lastname: Zhang
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Pons
authors.affiliation: the Department of Biology, Indiana University, Bloomington, Indiana 47405.
authors.forename: Melissa
authors.initials: M
authors.lastname: Konkol
authors.affiliation: the Department of Biology, Indiana University, Bloomington, Indiana 47405.
authors.forename: Daniel B
authors.initials: DB
authors.lastname: Kearns
authors.affiliation: the Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada, and the Program in Molecular Structure and Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
authors.forename: Dustin J
authors.initials: DJ
authors.lastname: Little
authors.affiliation: the Department of Biochemistry, University of Toronto, Toronto, Ontario M5S 1A8, Canada, and the Program in Molecular Structure and Function, Research Institute, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
authors.forename: P Lynne
authors.initials: PL
authors.lastname: Howell
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and dskurnik@rics.bwh.harvard.edu.
authors.forename: David
authors.initials: D
authors.lastname: Skurnik
authors.affiliation: From the Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, and gpier@rics.bwh.harvard.edu.
authors.forename: Gerald B
authors.initials: GB
authors.lastname: Pier
mesh_terms: D000117:Acetylglucosamine / Q000737:chemistry / Q000502:physiology*; D000907:Antibodies, Bacterial / Q000502:physiology; D001412:Bacillus subtilis / Q000502:physiology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000502:physiology; D018441:Biofilms; D053898:Biosynthetic Pathways; D004926:Escherichia coli; D018922:HL-60 Cells; D006801:Humans; D008958:Models, Molecular; D009895:Opsonin Proteins / Q000502:physiology; D010587:Phagocytosis; D011135:Polysaccharides, Bacterial; D017434:Protein Structure, Tertiary
chemical_list: D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D009895:Opsonin Proteins; D011135:Polysaccharides, Bacterial; C113831:poly-N-acetyl glucosamine; D000117:Acetylglucosamine
keywords: bacteria; bacterial adhesion; bacterial genetics; carbohydrate biosynthesis; carbohydrate structure; glycobiology; glycosyltransferase; protein expression; protein structure