Abstract
Cell wall polysaccharides (CWPS), which are usually covalently bound to the peptidoglycan and are closely associated with the cell wall, are considered as ubiquitous components of the cell envelope of gram-positive bacteria and play an important role as mediators of bacterial interactions with the environment. Here, we describe a simple method for purifying CWPS by extraction of bacterial cells with consecutive acid treatments. Purified CWPS are obtained by gel-filtration chromatography following treatment with HF. We also provide the methodology to easily assess the integrity of CWPS using high-resolution magic-angle spinning (HR-MAS) NMR.
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References
Delcour J, Ferain T, Deghorain M et al (1999) The biosynthesis and functionality of the cell-wall of lactic acid bacteria. In: Konings WN, Kuipers OP, In’t Veld JHJH (eds) Lactic acid bacteria: genetics, metabolism and applications. Springer, Dordrecht, pp 159–184
Kolenbrander PE, London J (1993) Adhere today, here tomorrow: oral bacterial adherence. J Bacteriol 175:3247–3252
Vinogradov E, Valence F, Maes E et al (2013) Structural studies of the cell wall polysaccharides from three strains of Lactobacillus helveticus with different autolytic properties: DPC4571, BROI, and LH1. Carbohydr Res 379:7–12
Cornelissen A, Sadovskaya I, Vinogradov E et al (2016) The baseplate of Lactobacillus delbrueckii bacteriophage Ld17 harbors a glycerophosphodiesterase. J Biol Chem 291:16816–16827
Vinogradov E, Sadovskaya I, Cornelissen A et al (2015) Structural investigation of cell wall polysaccharides of Lactobacillus delbrueckii subsp. bulgaricus 17. Carbohydr Res 413:93–99
Chapot-Chartier M-P, Vinogradov E, Sadovskaya I et al (2010) Cell surface of Lactococcus lactis is covered by a protective polysaccharide pellicle. J Biol Chem 285:10464–10471
Sadovskaya I, Vinogradov E, Courtin P et al (2017) Another brick in the wall: a rhamnan polysaccharide trapped inside peptidoglycan of Lactococcus lactis. mBio 8:e01303–e01317
Ainsworth S, Sadovskaya I, Vinogradov E et al (2014) Differences in lactococcal cell wall polysaccharide structure are major determining factors in bacteriophage sensitivity. mBio 5:e00880–14
Farenc C, Spinelli S, Vinogradov E et al (2014) Molecular insights on the recognition of a Lactococcus lactis cell wall pellicle by the phage 1358 receptor binding protein. J Virol 88:7005–7015
Chapot-Chartier M-P, Kulakauskas S (2014) Cell wall structure and function in lactic acid bacteria. Microb Cell Factories 13:S9
Prakobphol A, Linzer R (1980) Purification and immunological characterization of a rhamnose-glucose antigen from Streptococcus mutans 6517-T2 (serotype g). Infect Immun 30:140–146
Vinogradov E, Sadovskaya I, Grard T et al (2016) Structural studies of the rhamnose-rich cell wall polysaccharide of Lactobacillus casei BL23. Carbohydr Res 435:156–161
Li W, Lee REB, Lee RE, Li J (2005) Methods for acquisition and assignment of multidimensional high-resolution magic angle spinning NMR of whole cell bacteria. Anal Chem 77:5785–5792
Maes E, Mille C, Trivelli X et al (2009) Molecular phenotyping of mannosyltransferases-deficient Candida albicans cells by high-resolution magic angle spinning NMR. J Biochem (Tokyo) 145:413–419
Candela T, Maes E, Garénaux E et al (2011) Environmental and biofilm-dependent changes in a Bacillus cereus secondary cell wall polysaccharide. J Biol Chem 286:31250–31262
Dubois M, Gilles K, Hamilton JK et al (1951) A colorimetric method for the determination of sugars. Nature 168:167
Signoretto C, Lleò MM, Tafi MC et al (2000) Cell wall chemical composition of Enterococcus faecalis in the viable but nonculturable state. Appl Environ Microbiol 66:1953–1959
Brauge T, Sadovskaya I, Faille C et al (2016) Teichoic acid is the major polysaccharide present in the Listeria monocytogenes biofilm matrix. FEMS Microbiol Lett 363:fnv229
Hanoulle X, Wieruszeski J-M, Rousselot-Pailley P et al (2005) Monitoring of the ethionamide pro-drug activation in mycobacteria by (1)H high resolution magic angle spinning NMR. Biochem Biophys Res Commun 331:452–458
Acknowledgments
We acknowledge Douwe VanSideren (School of Microbiology, University College Cork, Ireland) and Marie-Pierre Chapot Chartier (Micalis Institute, INRA, France) for the use of Lactococcus lactis MG1363 and SK11. This work was supported by French ANR project “Lactophages” (ANR-11-BSV8-004-01).
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Sadovskaya, I., Guérardel, Y. (2019). Simple Protocol to Purify Cell Wall Polysaccharide from Gram-Positive Bacteria and Assess Its Structural Integrity. In: Brockhausen, I. (eds) Bacterial Polysaccharides. Methods in Molecular Biology, vol 1954. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9154-9_4
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DOI: https://doi.org/10.1007/978-1-4939-9154-9_4
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