New lactate- and pyruvate-containing polysaccharide and rhamnomannan with xylose residues from the cell wall of Rathayibacter oskolensis VKM Ac-2121T.

The cell wall of endophytic strain Rathayibacter oskolensis VKM Ac-2121T (family Microbacteriaceae, class Actinomycetes) was found to contain neutral and acidic glycopolymers. The neutral polymer is a block-type rhamnomannan partially should be substitutied by xylose residues, [→2)-α-[ß-D-Xylp-(1 â†’ 3)]-D-Manp-(1 â†’ 3)-α-D-Rhap-(1→]∼30 [→2)-α-D-Manp-(1 â†’ 3)-α-D-Rhap-(1→]∼45. The acidic polymer has branched chain, bearing lactate and pyruvate residues, →4)-α-D-[S-Lac-(2-3)-α-L-Rhap-(1 â†’ 3)]-D-Manp-(1 â†’ 3)-α-D-[4,6-R-Pyr]-D-Galp-(1 â†’ 3)-ß-D-Glcp-(1 â†’. The structures of both glycopolymers were not described in the Gram-positive bacteria to date. The glycopolymers were studied by chemical and NMR spectroscopic methods. The results of this study provide new data on diversity of bacterial glycopolymers and may prove useful in the taxonomy of the genus Rathayibacter and for understanding the molecular mechanisms of interaction between plants and plant endophytes.

A highly branched novel galactofuranan in the cell wall of Clavibacter tesselarius VKM Ac-1406T.

The structures of two cell wall glycopolymers were studied in the plant pathogenic bacterium Clavibacter tesselarius VKM Ac-1406T (family Microbacteriaceae, order Micrococcales, class Actinomycetes). The predominant polymer was a novel (1 â†’ 6)-linked ß-d-galactofuranan with a highly branched repeating unit, α-L-Rhap-(1 â†’ 3)-α-D-Galp-(1 â†’ 2)-[α-L-Rhap-(1 â†’ 3)]-α-D-Fucp-(1 →, at O-2 on every second galactofuranose residue. The second polymer present in small amounts was acidic with the repeating unit, →3)-α-D-Galp-(1 â†’ 3)-α-D-[4,6-S-Pyr]-Manp-(1 â†’ 3)-α-D-Manp-[2OAc]0.2-(1→, and was reported in all Clavibacter species investigated to date. The presented results expand our knowledges of structural diversity of phosphate-free cell wall glycopolymers and provide evidence in support of their taxonomic specificity for bacterial species and genera.

A novel cell wall galactofuranan in Clavibacter phaseoli VKM Ac-2641T.

A glycopolymer of novel structure was found in the cell wall of plant pathogen Clavibacter phaseoli VKM Ac-2641T (family Microbacteriaceae, class Actinomycetes). The glycopolymer was (1 â†’ 6)-linked ß-d-galactofuranan with side branched trisaccharide, α-D-Manp-(1 â†’ 2)-[α-D-Manp-(1 â†’ 3)]-α-D-Ribf-(1→ at O-2 on every second galactofuranose residue. The galactofuranan structure was established by chemical and NMR spectroscopic methods using one- and two-dimensional techniques 1H,1H COSY, TOCSY, ROESY and 1H,13C HSQC, HMBC. The results of this study provide new data on diversity of bacterial glycopolymers, may prove useful for bacterial taxonomy and contribute to the understanding of the host plant-microbiota interaction mechanisms.

Cell wall galactofuranan and pyruvate-containing galactomannan in the cell walls of Clavibacter strains.

The cell wall glycopolymer structures of plant-associated strains Clavibacter sp. VKM Ac-1371, Clavibacter sp. VKM Ac-1372 and Clavibacter sp. VKM Ac-1374, members of three putative new species (family Microbacteriaceae, class Actinobacteria) were studied. Each strain was found to contain two glycopolymers, neutral and acidic ones. The main chain of neutral polymer, identical in all three strains, is (1 â†’ 6)-linked ß-d-galactofuranan with every second galactofuranose residue substituted at position 2 by side disaccharide, α-d-Manp-(1 â†’ 2)-α-d-Ribf-(1 â†’ . The second, acidic polymer, is pyruvate-containing galactomannan with the repeating unit, →3)-α-d-Galp-(1 â†’ 3)-α-d-[4,6-S-Pyr]-Manp-(1 â†’ 3)-α-d-Manp-(1 â†’ . Reducing mannopyranose residues of the acidic polysaccharides repeating unit from strains VKM Ac-1372 and VKM Ac-1374 bear O-acetyl residues additionally. The cell wall glycopolymer structures were established by chemical and NMR spectroscopic methods with using one- and two-dimensional techniques 1H,1H COSY, TOCSY, ROESY and 1H,13C HSQC, HMBC. The results obtained provide new data on diversity of the bacterial cell wall glycopolymers and may prove valuable for microbial taxonomy and insight into the molecular mechanisms of interactions between bacteria and plants and also of bacterial adaptation to survival in desert systems.

D-Rhamnan and Pyruvate-Containing Teichuronic Acid from the Cell Wall of Rathayibacter sp. VKM Ac-2759.

Rathayibacter sp. VKM Ac-2759 (family Microbacteriaceae, class Actinobacteria) contains two glycopolymers in the cell wall. The main chain of rhamnan, glycopolymer 1, is built from the repeating tetrasaccharide units carrying terminal arabinofuranose residues at the non-reducing end, →3)-α-[α-D-Araf-(1→2)]-D-Rhap-(1→2)-α-D-Rhap-(1→3)-α-D-Rhap-(1→2)-α-D-Rhap-(1→. Similar to other described Rathayibacter species, rhamnose in the neutral glycopolymer of the VKM Ac-2759 strain is present in the D-configuration. Acetalated with pyruvic acid teichuronic acid, glycopolymer 2, is composed of the repeating tetrasaccharide units, →4)-ß-D-GlcpA-(1→4)-ß-D-Galp-(1→4)-ß-D-Glcp-(1→3)-ß-[4,6-S-Pyr]-D-Manp-(1→. Glycopolymers 1 and 2 were identified in prokaryotic microorganisms for the first time and their structures were established by chemical analysis and NMR spectroscopy. The obtained data can be used in taxonomic research, as well as for elucidating the mechanisms of plant colonization and infection by bacteria of the Rathayibacter genus.

Novel galactofuranan and pyruvylated galactomannan in the cell wall of Clavibacter michiganensis subsp. michiganensis VKM Ac-1403T.

The cell wall of Clavibacter michiganensis subsp. michiganensis VKM Ас-1403Т (family Microbacteriaceae, class Actinobacteria) contains two polysaccharides. The first one is neutral (1 â†’ 6) linked galactofuranan in which every second galactofuranose residue in the main chain substituted at position 3 by side trisaccharide, ß-D-GlcpNAc-(1 â†’ 3)-α-L-Rhap-(1 â†’ 2)-α-D-Fucp-(1 â†’. The second polymer is pyruvylated galactomannan with the repeating unit, →3)-α-D-Galp-(1 â†’ 3)-α-D-[4,6-S-Pyr]-Manp-(1 â†’ 3)-α-D-Manp-(1 â†’. The cell wall glycopolymer structures were established by chemical and NMR spectroscopic methods. The obtained results provide new data on the cell wall composition of plant pathogenic species of the genus Clavibacter and can promote understanding the molecular mechanisms involved in colonization and infection of plants.

D-rhamnan and teichuronic acid from the cell wall of Rathayibacter caricis VKM Ac-1799T.

The cell wall of Rathayibacter caricis VKM Ac-1799T (family Microbacteriaceae, class Actinobacteria) was found to contain both neutral and acidic glycopolymers. The first one is D-rhamnopyranan with main chain →2)-α-D-Rhap-(1 â†’ 3)-α-D-Rhap-(1→, where a part of 2-substituted residues bears as a side-chain at position 3 α-D-Manp residues or disaccharides α-D-Araf-(1→2)-α-D-Manp-(1 â†’ . The second polymer is a teichuronic acid with a branched repeating units composed of seven monosaccharides →4)-α-[ß-D-Manp-(1 â†’ 3)]-D-Glcp-(1 â†’ 4)-ß-D-GlcpA-(1 â†’ 2)-ß-[4,6Pyr]-D-Manp-(1 â†’ 4)-ß-L-Rhap-(1 â†’ 4)-ß-D-Glcp-(1 â†’ 4)-ß-D-Glcp-(1 â†’ . The structures of the polymers were determined by chemical and NMR spectroscopic methods.

New glycopolymers containing both D- and L-rhamnopyranoses from Rathayibacter iranicus VKM Ac-1602T cell wall.

The cell wall of Rathayibacter iranicus VKM Ac-1602T (family Microbacteriaceae, class Actinobacteria) is characterised by the absence of phosphate-containing and by the presence of two rhamnose-containing glycopolymers. The first is a branched rhamnomannan, in which 60% of mannose residues of the main chain are glycosylated by terminal mannose residues: →2)-α-D-Rhap-(1 → 3)-α-[α-D-Manp-(1 → 6)]-D-Manp-(1 → . The second is a branched teichuronic acid, in which all the rhamnose residues of the main chain are glycosylated by glucose residues:→3)-α-[α-D-Glcp-(1 → 2)]-L-Rhap-(1 → 4)-ß-D-GlcpA-(1 → 2)-α-D-Manp-(1 → 3)-α-D-Galp-(1 → 3)-ß-D-Glcp-(1 → . Both glycopolymers have the unique structures and described in the cell walls of Gram-positive bacteria for the first time. The obtained data allow for a more complete characterisation of the cell wall of the microorganism under investigation and can serve as a phenotypic characterisation of this bacterium. The glycopolymer structures were established using chemical and nuclear magnetic resonance (NMR) spectroscopy methods.

Pyruvylated cell wall glycopolymers of Promicromonospora citrea VKM A≿-665T and Promicromonospora sp. VKM A≿-1028.

The cell walls of two strains of the genus Promicromonospora (phylum Actinobacteria) were found to include non-phosphorylated anionic glycopolymers with pyruvic acid acetals of R-configuration. The cell wall of the type strain P. citrea 665T contains two glycopolymers of the sort, including the Kdn-teichulosonic acid with the repeating unit →6)-α-d-Gl≿p/→6)-α-d-Gl≿p3SO3--(1 â†’ 4)-α-[7,9Pyr]-Kdn-(2→, and the galactan with the repeating unit →3)-α-[4,6Pyr]-d-Galp-2OAc-(1 â†’ . The cell wall of Promicromonospora sp.VKM Ac-1028 contains the teichuronic acid with the repeating unit →6)-α-d-Gl≿p-(1 â†’ 4)-ß-[2,3Pyr]-d-GlcpA-(1 â†’ . The detected glycopolymer structures are reported for the first time. Presented results expand the notion on the diversity of the organic world and on the role of the structures and composition of cell wall polymers in bacterial taxonomy. The glycopolymer structures were established by using a combination of chemical methods, NMR- and IR-spectroscopy, and ESI MS.

Cell wall glycopolymers of Streptomyces albus, Streptomyces albidoflavus and Streptomyces pathocidini.

The cell wall glycopolymers of three strains of Streptomyces albus and the type strain of Streptomyces pathocidini were investigated. The structures of the glycopolymers were established using a combination of chemical and NMR spectroscopic methods. The cell wall of S. albus subsp. albus VKM Ac-35(T) was found to be comprised of three glycopolymers, viz. unsubstituted 1,5-poly(ribitol phosphate), 1,3-poly(glycerol phosphate) substituted with ß-D-glucopyranose, and the major polymer, a 3-deoxy-D-glycero-D-galacto-non-2-ulosonic acid (Kdn)-teichulosonic acid: ß-D-Glcp-(1 â†’ 8)-α-Kdnp-(2[(→6)-ß-D-Glcp-(1 â†’ 8)-α-Kdnp-(2 â†’] n 6)-ß-D-Glcp-(1 â†’ 8)-ß-Kdnp-(2-OH, where n ≥ 3. The cell walls of 'S. albus' J1074 and 'S. albus' R1-100 were found to contain three glycopolymers of identical structures, viz. unsubstituted 1,3- and 2,3-poly(glycerol phosphates), and the major polymer, a Kdn-teichulosonic acid with an unusual structure that has not been previously described: ß-D-Galp-(1 â†’ 9)-α-Kdnp-(2[(→3)-ß-D-Galp-(1 â†’ 9)-α-Kdnp-(2 â†’] n 3)-ß-D-Galp-(1 â†’ 9)-ß-Kdnp-(2-OH, where n ~ 7-8. The cell wall of S. pathocidini (formerly S. albus subsp. pathocidicus) VKM Ac-598(T) was found to contain two glycopolymers, viz. 1,3-poly(glycerol phosphate) partially O-glycosylated with 2-acetamido-2-deoxy-α-D-glucopyranose and/or O-acylated with L-lysine, and a poly(diglycosyl 1-phosphate) of hitherto unknown structure: -6)-α-D-Glcp-(1 â†’ 6)-α-D-GlcpNAc-(1-P-.

New structures and composition of cell wall teichoic acids from Nocardiopsis synnemataformans, Nocardiopsis halotolerans, Nocardiopsis composta and Nocardiopsis metallicus: a chemotaxonomic value.

The structures of the cell wall teichoic acids (TA) from some species of the genus Nocardiopsis were established by chemical and NMR spectroscopic methods. The cell walls of Nocardiopsis synnemataformans VKM Ac-2518(T) and Nocardiopsis halotolerans VKM Ac-2519(T) both contain two TA with unique structures-poly(polyol phosphate-glycosylpolyol phosphate)-belonging to the type IV TA. In both organisms, the minor TA have identical structures: poly(glycerol phosphate-N-acetyl-ß-galactosaminylglycerol phosphate) with the phosphodiester bond between C-3 of glycerol and C-4 of the amino sugar. This structure is found for the first time. The major TA of N. halotolerans has a hitherto unknown structure: poly(glycerol phosphate-N-acetyl-ß-galactosaminylglycerol phosphate), the N-acetyl-ß-galactosamine being acetalated with pyruvic acid at positions 4 and 6. The major TA of N. synnemataformans is a poly(glycerol phosphate-N-acetyl-ß-galactosaminylglycerol phosphate) with the phosphodiester bond between C-3 of glycerol and C-3 of the amino sugar. The cell walls of Nocardiopsis composta VKM Ac-2520 and N. composta VKM Ac-2521(T) contain only one TA, namely 1,3-poly(glycerol phosphate) partially substituted with N-acetyl-α-glucosamine. The cell wall of Nocardiopsis metallicus VKM Ac-2522(T) contains two TA. The major TA is 1,5-poly(ribitol phosphate), each ribitol unit carrying a pyruvate ketal group at positions 2 and 4. The structure of the minor TA is the same as that of N. composta. The results presented correlate well with the phylogenetic grouping of strains and confirm the species and strain specific features of cell wall TA in members of the genus Nocardiopsis.

A novel teichuronic acid, the major polymer from the cell wall of Actinoplanes lobatus VKM Ac-676(T.).

The cell wall of Actinoplanes lobatus VKM Ac-676(T) (family Micromonosporaceae; order Actinomycetales; class Actinobacteria) contains two anionic glycopolymers. The major one is a teichuronic acid with a linear structure of the chain and heterogeneous repeating units, consisting of glucopyranose residues and statistically alternating residues of diaminouronic acids having the d-manno,l-gulo and d-gluco configurations in a ratio of 6:3:1, respectively. The teichuronic acid of this structure has not hitherto been found in Gram-positive bacteria. The minor glycopolymer is a teichoic acid of poly(glycosylpolyol phosphate) nature with the repeating unit -6)-[α-d-Manp(1→4)]-ß-d-Galp-(1→2)-snGro-(3-P-. The structures of the glycopolymers were established by chemical methods and NMR spectroscopy.

Novel teichulosonic acid from cell wall of Streptomyces coelicolor M145.

The cell wall of Streptomyces coelicolor M145, a prototrophic plasmidless (SCP1(-) SCP2(-)) variant of strain S. coelicolor A3(2) contains the main glycopolymer represented by Kdn-containing teichulosonic acid with unusual structure which has not been described so far: [structure: see text]. The minor polymer was found to be a poly(diglycosyl 1-phosphate) with the following repeating unit: -6)-α-Galp-(1→6)-α-GlcpNAc-(1-P-. The structures of both glycopolymers were established by using a combination of chemical and NMR spectroscopic methods.

Novel teichulosonic acid from cell walls of some representatives of the genus Kribbella.

Cell walls of each of five bacterial strains belonging to the genus Kribbella (family Nocardioidaceae, order Actinomycetales) contain a neutral polysaccharide (mannan) and teichulosonic acid of novel structure in different proportions. The novel teichulosonic acid found in strains VKM Ac-2500, VKM Ас-2568, VKM Ас-2572, and VKM Ас-2575 is a heteropolymer with an irregular structure where fragments I (predominant) alternate with fragments II (minor): The teichulosonic acid from Kribbella sp. VKM Ac-2527 has in general a structure similar to that above with the exception that the Pse residue is randomly glycosylated at O-4 with ß-l-Rhap (along with α-d-Galp3OMe or α-d-Galp2,3OMe). The strain VKM Ac-2572 contained additionally teichuronic acid with the disaccharide repeating unit consisted of aminomannuronic acid and 2,3-diacetamido-2,3-dideoxy-α-glucopyranose. The mannan, a polysaccharide common to all of the strains, is built of (1→6)-linked α-d-mannopyranose substituted with α-d-mannopyranose at O-2. The structures of all the glycopolymers were established by a combination of chemical and NMR spectroscopic methods.

New cell wall glycopolymers of the representatives of the genus Kribbella.

Each of the cell walls of four representatives of the genus Kribbella (order Actinomycetales; suborder Propionibacterineae; family Nocardioidaceae) contains a neutral polysaccharide and an acidic polysaccharide with unusual structures. Common to all four strains studied is a mannan with the following repeating unit: [structure: see text]. In the cell wall of the strain VKM Ac-2541, a teichulosonic acid was identified with a monosaccharide component that has not hitherto been found in Gram-positive bacteria, viz., pseudaminic acid, and an unusual linkage type in the polymeric chain, [structure: see text] where R=capital EN, Cyrillic (45%), alpha-D-Galp3OMe (37%) or alpha-D-Galp2,3OMe (18%). The anionic cell wall components of three other strains are represented by teichuronic acids with a rare constituent, viz., a diaminosugar, 2,3-diacetamido-2,3-dideoxyglucopyranose. The structures of their repeating units differ in the nature of the acidic components: -->4)-beta-D-Manp2,3NAcA-(1-->6)-alpha-D-Glcp2,3NAc-(1--> (VKM capital A, Cyrillicsmall es, Cyrillic-2538 and VKM capital A, Cyrillicsmall es, Cyrillic-2540) and -->4)-beta-D-ManpNAcA-(1-->6)-alpha-D-Glcp2,3NAc-(1--> (VKM capital A, Cyrillicsmall es, Cyrillic-2539). The structures of all the glycopolymers were established by chemical and NMR spectroscopic methods; they are identified in Gram-positive bacteria for the first time.

A new neutral polymer from the cell wall of actinomycete Kineosporia aurantiaca VKM Ac-702T.

The major cell wall polymer of Kineosporia aurantiaca VKM Ac-702T a representative of the suborder Frankineae, is a galactomannan with a repeating unit of the following structure: -->3)-beta-D-Galp-(1-->6)-beta-D-Manp-(1-->4)-beta-D-Manp-(1-->3)-beta-D-Galp-(1-->4)-beta-D-Manp-(1-->4)-beta-D-Manp-(1--> that has not been reported so far. This was established using chemical degradation methods and NMR spectroscopy. The polysaccharide identified in the present study provides the second example of neutral galactomannans in actinomycete cell walls. The cell wall of K. aurantiaca VKM Ac-702T also contains a minor teichoic acid, viz., 1,3-poly(glycerol phosphate) partially substituted with alpha-glucosamine residues, only part of which are N-acetylated.

Cell wall anionic polymers of Streptomyces sp. MB-8, the causative agent of potato scab.

The cell wall of Streptomyces sp. MB-8 contains a major teichoic acid, viz., 1,3-poly(glycerol phosphate) substituted with N-acetyl-alpha-D-glucosamine (the degree of substitution is 60%), a minor teichoic acid, viz., non-substituted poly(glycerol phosphate), and a family of Kdn (3-deoxy-D-glycero-D-galacto-non-2-ulopyranosonic acid)-containing oligomers of the following general structure: [carbohydrate structure: see text]. The composition of the oligomers was established using MALDI-TOF mass spectroscopy. The present study provides the second example of the identification of Kdn as a component of cell wall polymers of streptomycetes, which are the causative agents of potato scab.

Novel cell-wall teichoic acid from Nocardiopsis albus subsp. albus as a species-specific marker.

Cell walls of the three Nocardiopsis albus subsp. albus strains, DSM 43377T, 43378 and 43120 contain structurally identical teichoic acids. The cell wall of each strain has three distinct types of teichoic acids: (1) unsubstituted 3,5-poly(ribitol phosphate), (2) 1,3-poly(glycerol phosphate) partially substituted at C-2 with α-N-acetylglucosamine residues, and (3) 1,5-poly(ribitol phosphate) with each ribitol unit carrying a 2,4-pyruvate ketal group. Types 1 and 3 are reported in prokaryotes for the first time. The structure of the teichoic acids was elucidated by chemical analysis and NMR-spectroscopic methods. Structural identity of the teichoic acids from the three strains belonging to the same species may demonstrate the species-specificity of these polymers.