title: Structural analysis of a novel anionic polysaccharide from Porphyromonas gingivalis strain W50 related to Arg-gingipain glycans.
abstract: The Arg-gingipains (RgpsA and B) of Porphyromonas gingivalis are a family of extracellular cysteine proteases and are important virulence determinants of this periodontal bacterium. A monoclonal antibody, MAb1B5, which recognizes an epitope on glycosylated monomeric RgpAs also cross-reacts with a cell-surface polysaccharide of P. gingivalis W50 suggesting that the maturation pathway of the Arg-gingipains may be linked to the biosynthesis of a surface carbohydrate. We report the purification and structural characterization of the cross-reacting anionic polysaccharide (APS), which is distinct from both the lipopolysaccharide and serotype capsule polysaccharide of P. gingivalis W50. The structure of APS was determined by 1D and 2D NMR spectroscopy and methylation analysis, which showed it to be a phosphorylated branched mannan. The backbone is built up of alpha-1,6-linked mannose residues and the side-chains contain alpha-1,2-linked mannose oligosaccharides of different lengths (one to two sugar residues) attached to the backbone via 1,2-linkage. One of the side-chains in the repeating unit contains Manalpha1-2Manalpha1-phosphate linked via phosphorus to a backbone mannose at position 2. De-O-phosphorylation of APS abolished cross-reactivity suggesting that Manalpha1-2Manalpha1-phosphate fragment forms part of the epitope recognized by MAb1B5. This phosphorylated branched mannan represents a novel polysaccharide that is immunologically related to the post-translational additions of Arg-gingipains.
authors.affiliation: MRC Molecular Pathogenesis Group, Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts and The London, Queen Mary's School of Medicine and Dentistry, 4 Newark Street, London E1 2AT, UK.
authors.forename: Nikolay
authors.initials: N
authors.lastname: Paramonov
authors.forename: Minnie
authors.initials: M
authors.lastname: Rangarajan
authors.forename: Ahmed
authors.initials: A
authors.lastname: Hashim
authors.forename: Alex
authors.initials: A
authors.lastname: Gallagher
authors.forename: Joseph
authors.initials: J
authors.lastname: Aduse-Opoku
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Slaney
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Hounsell
authors.forename: Michael A
authors.initials: MA
authors.lastname: Curtis
mesh_terms: D018829:Adhesins, Bacterial / Q000737:chemistry*; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D003546:Cysteine Endopeptidases / Q000737:chemistry*; D000080867:Gingipain Cysteine Endopeptidases; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D019906:Nuclear Magnetic Resonance, Biomolecular; D009844:Oligosaccharides / Q000737:chemistry*; D016966:Porphyromonas gingivalis / Q000737:chemistry* / Q000648:ultrastructure; D012703:Serotyping
chemical_list: D018829:Adhesins, Bacterial; D000080867:Gingipain Cysteine Endopeptidases; D009844:Oligosaccharides; D003546:Cysteine Endopeptidases