Abstract
Spirochaeta aurantia is a free-living saprophytic spirochete that grows easily in simple laboratory media, and thus can be used as a model for the investigation of surface carbohydrate structures in spirochetae, which are normally not available in sufficient amounts. Freeze-substitution electron microscopy indicated the presence of a capsule-like material projecting from the surface of S. aurantia. Extraction of cells gave two major glycolipids, the one with a higher molecular mass glycolipid was designated large glycolipid A (LGLA). LGLA contained small amount of branched and unsaturated O-linked fatty acids, l-rhamnose, l-fucose, d-xylose, d-mannose, d-glucosamine, d-glycero-d-gluco-heptose (DDglcHep), d-glycero-d-manno-heptose (DDHep), and a novel branched tetradeoxydecose monosaccharide, which we proposed to call aurantose (Aur). The carbohydrate structure of LGLA was extremely complex and consisted of the repeating units built of 11 monosaccharides, arrangement of nine of them was determined as:
which wasdeduced from the NMR and chemical data on the LGLA and its fragments, obtained by various degradations. Tentative position of two remaining sugars is proposed. LGLA was negative for gelation of Limulus amebocyte lysate, did not contain lipid A, and was unable to activate any known Toll-like receptors.
Similar content being viewed by others
References
Canale-Parola, E.: Spirochaeta. In: Krieg, N.R. (ed.) Bergey’s Manual of Systematic Bacteriology. Williams and Wilkins, Baltimore, MD (1984)
Raetz, C.R., Whitfield, C.: Lipopolysaccharide endotoxins. Annu. Rev. Biochem. 71, 635–700 (2002). doi:10.1146/annurev.biochem.71.110601.135414
Triantafilou, M., Lepper, P.M., Briault, C.D., Ahmed, M.A., Dmochowski, J.M., Schumann, C., Triantafilou, K.: Chemokine receptor 4 (CXCR4) is part of the lipopolysaccharide “sensing apparatus”. Eur. J. Immunol. 38, 192–203 (2008). doi:10.1002/eji.200636821
Triantafilou, M., Triantafilou, K.: The dynamics of LPS recognition: complex orchestration of multiple receptors. J. Endotoxin Res. 11, 5–11 (2005)
Que-Gewirth, N.L., Ribeiro, A.A., Kalb, S.R., Cotter, R.J., Bulach, D.M., Adler, B., Girons, I.S., Werts, C., Raetz, C.R.: A methylated phosphate group and four amide-linked acyl chains in Leptospira interrogans lipid A. The membrane anchor of an unusual lipopolysaccharide that activates TLR2. J. Biol. Chem. 279, 25420–25429 (2004). doi:10.1074/jbc.M400598200
Fraser, C.M., Casjens, S., Huang, W.M., Sutton, G.G., Clayton, R., Lathigra, R., White, O., Ketchum, K.A., Dodson, R., Hickey, E.K., Gwinn, M., Dougherty, B., Tomb, J.F., Fleischmann, R.D., Richardson, D., Peterson, J., Kerlavage, A.R., Quackenbush, J., Salzberg, S., Hanson, M., van Vugt, R., Palmer, N., Adams, M.D., Gocayne, J., Venter, J.C.: Genomic sequence of a Lyme disease spirochaete, Borrelia burgdorferi. Nature 390, 580–586 (1997). doi:10.1038/37551
Ben-Menachem, G., Kubler-Kielb, J., Coxon, B., Yergey, A., Schneerson, R.: A newly discovered cholesteryl galactoside from Borrelia burgdorferi. Proc. Natl. Acad. Sci. U. S. A. 100, 7913–7918 (2003). doi:10.1073/pnas.1232451100
Schroder, N.W., Schombel, U., Heine, H., Gobel, U.B., Zahringer, U., Schumann, R.R.: Acylated cholesteryl galactoside as a novel immunogenic motif in Borrelia burgdorferi sensu stricto. J. Biol. Chem. 278, 33645–33653 (2003). doi:10.1074/jbc.M305799200
Schultz, C.P., Wolf, V., Lange, R., Mertens, E., Wecket, J., Naumann, D., Zahringer, U.: Evidence for a new type of outer membrane lipid in oral spirochete Treponema denticola: functioning permeation barrier without lipopolysaccharides. J. Biol. Chem. 273, 15661–15666 (1998). doi:10.1074/jbc.273.25.15661
Schroder, N.W., Opitz, B., Lamping, N., Michelsen, K.S., Zahringer, U., Gobel, U.B., Schumann, R.R.: Involvement of lipopolysaccharide binding protein, CD14, and Toll-like receptors in the initiation of innate immune responses by Treponema glycolipids. J. Immunol. 165, 2683–2693 (2000)
Kesavalu, L., Falk, C.W., Davis, K.J., Steffen, M.J., Xu, X., Holt, S.C., Ebersole, J.L.: Biological characterization of lipopolysaccharide from Treponema pectinovorum. Infect. Immun. 70, 211–217 (2002). doi:10.1128/IAI.70.1.211-217.2002
Breznak, J.A., Canale-Parola, E.: Morphology and physiology of Spirochaeta aurantia strains isolated from aquatic habitats. Arch. Microbiol. 105, 1–12 (1975). doi:10.1007/BF00447104
Lilburn, T.G., Kim, K.S., Ostrom, N.E., Byzek, K.R., Leadbetter, J.R., Breznak, J.A.: Nitrogen fixation by symbiotic and free-living spirochetes. Science 292, 2495–2498 (2001). doi:10.1126/science.1060281
Vinogradov, E., Paul, C.J., Li, J., Zhou, Y., Lyle, E.A., Tapping, R.I., Kropinski, A.M., Perry, M.B.: The structure and biological characteristics of the Spirochaeta aurantia outer membrane glycolipid LGLB. Eur. J. Biochem. 271, 4685–4695 (2004). doi:10.1111/j.1432-1033.2004.04433.x
Darveau, R.P., Hancock, R.E.: Procedure for isolation of bacterial lipopolysaccharides from both smooth and rough Pseudomonas aeruginosa and Salmonella typhimurium strains. J. Bacteriol. 155, 831–838 (1983)
Kropinski, A.M., Ghiorse, W.C., Greenberg, E.P.: The intracellular polyglucose storage granules of Spirochaeta aurantia. Arch. Microbiol. 184, 289–295 (1988). doi:10.1007/BF00407794
Feingold, D.S., Youngner, J.S., Chen, J.: Interferon production in mice by cell wall mutants of Salmonella typhimurium. III. Role of lipid moiety of bacterial lipopolysaccharide in Interferon production in animals. Ann.N.Y. Acad. Sci. 173, 249–254 (1970). doi:10.1111/j.1749-6632.1970.tb53415.x
Bock, K., Pedersen, C.: Carbon-13 nuclear magnetic resonance spectroscopy of monosaccharides. Adv. Carbohydr. Chem. Biochem. 41, 27–66 (1983). doi:10.1016/S0065-2318(08)60055-4
Lipkind, G.M., Shashkov, A.S., Knirel, Y.A., Vinogradov, E.V., Kochetkov, N.K.: A computer-assisted structural analysis of regular polysaccharides on the basis of 13C-n.m.r. data. Carbohydr. Res. 175, 59–75 (1988). doi:10.1016/0008-6215(88)80156-3
Hashimoto, M., Asai, Y., Jinno, T., Adachi, S., Kusumoto, S., Ogawa, T.: Structural elucidation of polysaccharide part of glycoconjugate from Treponema medium ATCC 700293. Eur. J. Biochem. 270, 2671–2679 (2003). doi:10.1046/j.1432-1033.2003.03644.x
Asai, Y., Ohyama, Y., Taiji, Y., Makimura, Y., Tamai, R., Hashimoto, M., Ogawa, T.: Treponema medium glycoconjugate inhibits activation of human gingival fibroblasts stimulated with phenol–water extracts of periodontopathic bacteria. J. Dent. Res. 84, 456–461 (2005). doi:10.1177/154405910508400511
Sambrook, J., Fritsch, E.F., Maniatis, T.: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, MA, USA (1989)
Lesse, A.J., Campagnari, A.A., Bittner, W.E., Apicella, M.A.: Increased resolution of lipopolysaccharides and liopoligosaccharides utilizing tricine–sodium dodecyl sulfate-polyacrylamide gel electrophoresis. J. Immunol. Methods 126, 107–109 (1990). doi:10.1016/0022-1759(90)90018-Q
Tsai, C.M., Frasch, C.E.: A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal. Biochem. 119, 115–119 (1982). doi:10.1016/0003-2697(82)90673-X
Dmitriev, B.A., Knirel, Y.A., Kochetkov, N.K.: Selective cleavage of glycosidic linkages: Studies with the model compound benzyl 2-acetamido-2-deoxy-6-O-a-d-mannopyranosyl-a-d-glucopyranoside. Carbohydr. Res. 30, 45–50 (1973). doi:10.1016/S0008-6215(00)82171-0
Tapping, R.I., Akashi, S., Miyake, K., Godowski, P.J., Tobias, P.S.: Toll-like receptor 4, but not toll-like receptor 2, is a signaling receptor for Escherichia and Salmonella lipopolysaccharides. J. Immunol. 165, 5780–5787 (2000)
Paul, T.R., Beveridge, T.J.: Ultrastructure of mycobacterial surfaces by freeze-substitution. Zentralbl. Bakteriol. 279, 450–457 (1993)
Graham, L.L., Harris, R., Villiger, W., Beveridge, T.J.: Freeze-substitution of Gram-negative eubacteria: general cell morphology and envelope profiles. J. Bacteriol. 173, 1623–1633 (1991)
Dubois, M., Gilles, K.A., Hamilton, J.K., Rebers, P.A., Smith, F.: Colorimetric method for determination of sugars and related substances. Anal. Chem. 28, 350–356 (1956). doi:10.1021/ac60111a017
Johnson, A.R.: Improved method of hexosamine determination. Anal. Biochem. 44, 628–635 (1971). doi:10.1016/0003-2697(71)90252-1
Karkhanis, Y.D., Zeltner, J.Y., Jackson, J.J., Carlo, D.J.: A new and improved microassay to determine 2-keto-3-deoxyoctonate in lipopolysaccharide of gram-negative bacteria. Anal. Biochem. 85, 595–601 (1978). doi:10.1016/0003-2697(78)90260-9
Holst, O.: In: Holst, O. (ed.) Methods in Molecular Biology, Bacterial Toxins: Methods and Protocols, pp. 345–353. Humana, Totowa, NJ (2000)
Leontein, K., Lindberg, B., Loenngren, J.: Assignment of absolute configuration of sugars by g.l.c. of their acetylated glycosides formed from chiral alcohols. Carbohydr. Res. 62, 359–362 (1978). doi:10.1016/S0008-6215(00)80882-4
Acknowledgements
This work was supported by a Natural Sciences and Engineering Research Council of Canada Discovery grant to A.M.K. Authors are very indebted to Dr. Cristina De Castro, Università di Napoli “Federico II", for the identification of paratose and to Ms. Diane Moyles at the University of Guelph for freeze substitution electron microscopy.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Paul, C.J., Lyle, E.A., Beveridge, T.J. et al. Characterization of the cell surface glycolipid from Spirochaeta aurantia . Glycoconj J 26, 1097–1108 (2009). https://doi.org/10.1007/s10719-009-9230-4
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10719-009-9230-4