Dertli E.Colquhoun I.J.Gunning A.P.Bongaerts R.J.Le Gall G.Bonev B.B.Mayer M.J.Narbad A.20.04.20192019-04-2020.04.20192019-04-2020130021-9258https://dx.doi.org/10.1074/jbc.M113.507418https://hdl.handle.net/20.500.12403/835Exopolysaccharides were isolated and purified from Lactobacillus johnsonii FI9785, which has previously been shown to act as a competitive exclusion agent to control Clostridium perfringens in poultry. Structural analysis by NMR spectroscopy revealed that L. johnsonii FI9785 can produce two types of exopolysaccharide: EPS-1 is a branched dextran with the unusual feature that every backbone residue is substituted with a 2-linked glucose unit, and EPS-2 was shown to have a repeating unit with the following structure: -6)-?-Glcp-(1-3)-?-Glcp-(1-5)-?-Galf-(1-6)-?-Glcp-(1-4) -?-Gal p-(1-4)-?-Glcp-(1-. Sites on both polysaccharides were partially occupied by substituent groups: 1-phosphoglycerol and O-acetyl groups in EPS-1 and a single O-acetyl group in EPS-2. Analysis of a deletion mutant (?epsE) lacking the putative priming glycosyltransferase gene located within a predicted eps gene cluster revealed that the mutant could produce EPS-1 but not EPS-2, indicating that epsE is essential for the biosynthesis of EPS-2. Atomic force microscopy confirmed the localization of galactose residues on the exterior of wild type cells and their absence in the ?epsE mutant. EPS2 was found to adopt a random coil structural conformation. Deletion of the entire 14-kb eps cluster resulted in an acapsular mutant phenotype that was not able to produce either EPS-2 or EPS-1. Alterations in the cell surface properties of the EPS-specific mutants were demonstrated by differences in binding of an anti-wild type L. johnsonii antibody. These findings provide insights into the biosynthesis and structures of novel exopolysaccharides produced by L. johnsonii FI9785, which are likely to play an important role in biofilm formation, protection against harsh environment of the gut, and colonization of the host. © 2013 by The American Society for Biochemistry and Molecular Biology, Inc.eninfo:eu-repo/semantics/openAccessBiofilm formationCell surface propertiesClostridium perfringensExopolysaccharidesGalactose residuesHarsh environmentPriming glycosyltransferase genesStructural conformationsAtomic force microscopyBiochemistryBiosynthesisCell membranesNuclear magnetic resonance spectroscopyPolysaccharidesMetabolitesbacterium antibodyexopolysaccharidegalactoseglycosyltransferasearticleatomic force microscopybacterial strainbacterium culturebiosynthesiscarbohydrate analysisconformationcontrolled studyflow cytometrygene clusterLactobacillus johnsoniiLactobacillus johnsonii FI9785mutantnonhumannuclear magnetic resonance spectroscopyphenotypepriority journaltransmission electron microscopywild typeAtomic Force MicroscopyBacteriaCarbohydrate Structureeps ClusterExopolysaccharideLactobacillus johnsoniiMutantNuclear Magnetic ResonanceBacterial ProteinsCarbohydrate ConformationGenes, BacterialGlucosyltransferasesLactobacillusMultigene FamilyMutationPolysaccharides, BacterialBiofilm formationCell surface propertiesClostridium perfringensExopolysaccharidesGalactose residuesHarsh environmentPriming glycosyltransferase genesStructural conformationsAtomic force microscopyBiochemistryBiosynthesisCell membranesNuclear magnetic resonance spectroscopyPolysaccharidesMetabolitesbacterium antibodyexopolysaccharidegalactoseglycosyltransferasearticleatomic force microscopybacterial strainbacterium culturebiosynthesiscarbohydrate analysisconformationcontrolled studyflow cytometrygene clusterLactobacillus johnsoniiLactobacillus johnsonii FI9785mutantnonhumannuclear magnetic resonance spectroscopyphenotypepriority journaltransmission electron microscopywild typeAtomic Force MicroscopyBacteriaCarbohydrate Structureeps ClusterExopolysaccharideLactobacillus johnsoniiMutantNuclear Magnetic ResonanceBacterial ProteinsCarbohydrate ConformationGenes, BacterialGlucosyltransferasesLactobacillusMultigene FamilyMutationPolysaccharides, BacterialArticle28844319383195110.1074/jbc.M113.507418240195312-s2.0-84887093174Q1WOS:000330596200046Q1