title: Characterization of a Novel d-Glycero-d-talo-oct-2-ulosonic acid-substituted Lipid A Moiety in the Lipopolysaccharide Produced by the Acetic Acid Bacterium Acetobacter pasteurianus NBRC 3283.
abstract: Acetobacter pasteurianus is an aerobic Gram-negative rod that is used in the fermentation process used to produce the traditional Japanese black rice vinegar kurozu. Previously, we found that a hydrophobic fraction derived from kurozu stimulates Toll-like receptors to produce cytokines. LPSs, particularly LPS from A. pasteurianus, are strong candidates for the immunostimulatory component of kurozu. The LPS of A. pasteurianus remains stable in acidic conditions during the 2 years of the abovementioned fermentation process. Thus, we hypothesized that its stability results from its structure. In this study, we isolated the LPS produced by A. pasteurianus NBRC 3283 bacterial cells and characterized the structure of its lipid A component. The lipid A moiety was obtained by standard weak acid hydrolysis of the LPS. However, the hydrolysis was incomplete because a certain proportion of the LPS contained acid-stable d-glycero-d-talo-oct-2-ulosonic acid (Ko) residues instead of the acid-labile 3-deoxy-d-manno-oct-2-ulosonic acid residues that are normally found in typical LPS. Even so, we obtained a Ko-substituted lipid A with a novel sugar backbone, α-Man(1-4)[α-Ko(2-6)]β-GlcN3N(1-6)α-GlcN(1-1)α-GlcA. Its reducing end GlcN(1-1)GlcA bond was also found to be quite acid-stable. Six fatty acids were attached to the backbone. Both the whole LPS and the lipid A moiety induced TNF-α production in murine cells via Toll-like receptor 4, although their activity was weaker than those of Escherichia coli LPS and lipid A. These results suggest that the structurally atypical A. pasteurianus lipid A found in this study remains stable and, hence, retains its immunostimulatory activity during acetic acid fermentation.
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan, hassy@eng.kagoshima-u.ac.jp.
authors.forename: Masahito
authors.initials: M
authors.lastname: Hashimoto
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan.
authors.forename: Mami
authors.initials: M
authors.lastname: Ozono
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan.
authors.forename: Maiko
authors.initials: M
authors.lastname: Furuyashiki
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan.
authors.forename: Risako
authors.initials: R
authors.lastname: Baba
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan.
authors.forename: Shuhei
authors.initials: S
authors.lastname: Hashiguchi
authors.affiliation: From the Department of Chemistry, Biotechnology, and Chemical Engineering, Kagoshima University, Kagoshima 890-0065, Japan.
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Suda
authors.affiliation: the Department of Chemistry, Graduate School of Science, Osaka University, Osaka 560-0043, Japan, and.
authors.forename: Koichi
authors.initials: K
authors.lastname: Fukase
authors.affiliation: the Faculty of Science and Technology, Keio University, Kanagawa 223-8522, Japan.
authors.forename: Yukari
authors.initials: Y
authors.lastname: Fujimoto
mesh_terms: D000091:Acetobacter / Q000737:chemistry* / Q000276:immunology; D000818:Animals; D002236:Carbohydrate Conformation; D002460:Cell Line; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008050:Lipid A / Q000737:chemistry* / Q000276:immunology; D051379:Mice; D051197:Toll-Like Receptor 4 / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
chemical_list: D008050:Lipid A; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha
keywords: d- glycero-d-talo-oct-2-ulosonic acid (Ko); glycoconjugate; glycolipid; lipid A; lipopolysaccharide (LPS); mass spectrometry (MS); nuclear magnetic resonance (NMR)