title: Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications.
abstract: Tannerella forsythia is an anaerobic, Gram-negative periodontal pathogen. A unique O-linked oligosaccharide decorates the bacterium's cell surface proteins and was shown to modulate the host immune response. In our study, we investigated the biosynthesis of the nonulosonic acid (NulO) present at the terminal position of this glycan. A bioinformatic analysis of T. forsythia genomes revealed a gene locus for the synthesis of pseudaminic acid (Pse) in the type strain ATCC 43037 while strains FDC 92A2 and UB4 possess a locus for the synthesis of legionaminic acid (Leg) instead. In contrast to the NulO in ATCC 43037, which has been previously identified as a Pse derivative (5-N-acetimidoyl-7-N-glyceroyl-3,5,7,9-tetradeoxy-l-glycero-l-manno-NulO), glycan analysis of strain UB4 performed in this study indicated a 350-Da, possibly N-glycolyl Leg (3,5,7,9-tetradeoxy-d-glycero-d-galacto-NulO) derivative with unknown C5,7 N-acyl moieties. We have expressed, purified and characterized enzymes of both NulO pathways to confirm these genes' functions. Using capillary electrophoresis (CE), CE-mass spectrometry and NMR spectroscopy, our studies revealed that Pse biosynthesis in ATCC 43037 essentially follows the UDP-sugar route described in Helicobacter pylori, while the pathway in strain FDC 92A2 corresponds to Leg biosynthesis in Campylobacter jejuni involving GDP-sugar intermediates. To demonstrate that the NulO biosynthesis enzymes are functional in vivo, we created knockout mutants resulting in glycans lacking the respective NulO. Compared to the wild-type strains, the mutants exhibited significantly reduced biofilm formation on mucin-coated surfaces, suggestive of their involvement in host-pathogen interactions or host survival. This study contributes to understanding possible biological roles of bacterial NulOs.
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Valentin
authors.initials: V
authors.lastname: Friedrich
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Bettina
authors.initials: B
authors.lastname: Janesch
authors.affiliation: Department of Chemistry, Universität für Bodenkultur Wien, Muthgasse 18, Vienna, Austria.
authors.forename: Markus
authors.initials: M
authors.lastname: Windwarder
authors.affiliation: Department of Chemistry, Universität für Bodenkultur Wien, Muthgasse 18, Vienna, Austria.
authors.forename: Daniel
authors.initials: D
authors.lastname: Maresch
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Matthias L
authors.initials: ML
authors.lastname: Braun
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Zoë A
authors.initials: ZA
authors.lastname: Megson
authors.affiliation: National Research Council, Human Health Therapeutics Portfolio, 100 Sussex Drive, Ottawa, ON, Canada.
authors.forename: Evgeny
authors.initials: E
authors.lastname: Vinogradov
authors.affiliation: National Research Council, Human Health Therapeutics Portfolio, 100 Sussex Drive, Ottawa, ON, Canada.
authors.forename: Marie-France
authors.initials: MF
authors.lastname: Goneau
authors.affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, 311 Foster Hall, 3435 Main St. Buffalo, New York, USA.
authors.forename: Ashu
authors.initials: A
authors.lastname: Sharma
authors.affiliation: Department of Chemistry, Universität für Bodenkultur Wien, Muthgasse 18, Vienna, Austria.
authors.forename: Friedrich
authors.initials: F
authors.lastname: Altmann
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Paul
authors.initials: P
authors.lastname: Messner
authors.affiliation: National Research Council, Human Health Therapeutics Portfolio, 100 Sussex Drive, Ottawa, ON, Canada.
authors.forename: Ian C
authors.initials: IC
authors.lastname: Schoenhofen
authors.affiliation: Department of NanoBiotechnology, NanoGlycobiology Unit, Universität für Bodenkultur Wien, Muthgasse 11, Vienna, Austria.
authors.forename: Christina
authors.initials: C
authors.lastname: Schäffer
mesh_terms: D053898:Biosynthetic Pathways / Q000235:genetics*; D016680:Genome, Bacterial / Q000235:genetics; D006031:Glycosylation; D054884:Host-Pathogen Interactions / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000235:genetics*; D009844:Oligosaccharides / Q000235:genetics / Q000378:metabolism; D012794:Sialic Acids / Q000096:biosynthesis; D013400:Sugar Acids / Q000378:metabolism; D000070436:Tannerella forsythia / Q000201:enzymology / Q000235:genetics* / Q000472:pathogenicity
chemical_list: C513444:5,7-diacetamido-3,5,7,9-tetradeoxynonulosonic acid; C110570:5,7-diacetamido-8-O-acetyl-3,5,7,9-tetradeoxy-glycero-talo-nonulosonic acid; D008565:Membrane Proteins; D009844:Oligosaccharides; D012794:Sialic Acids; D013400:Sugar Acids
keywords: bacterium; biofilm; biosynthesis pathway; pseudaminic and legionaminic acid