The Cell Wall Teichoic Acids of Streptomycetes from the “Streptomyces cyaneus” Cluster

The cell wall anionic polymers of the 13 species of the Streptomyces cyaneus cluster have a similar structure and contain -glucosylated 1,5-poly(ribitol phosphate) and 1,3-poly(glycerol phosphate). In the degree of glucosylation of the ribitol phosphate units of their teichoic acids, the cluster members can be divided into two groups. The streptomycetes of the first group (S. afghaniensis, S. janthinus, S. purpurascens, S. roseoviolaceus, and S. violatus) are characterized by a very similar structure of their cell walls, the completely glucosylated 1,5-poly(ribitol phosphate) chains, and a high degree of DNA homology (67–88% according to literature data). The cell wall teichoic acids of the second group (S. azureus, S. bellus, S. caelestis, S. coeruleorubidus, S. curacoi, and S. violarus) differ in the degree of -glucosylation of their 1,5-poly(ribitol phosphate) chains and have a lower level of DNA homology (54–76% according to literature data). Two streptomycetes of the cluster (S. cyaneus and S. hawaiiensis) are genetically distant from the other cluster members but have the same composition and structure of the cell wall teichoic acids as the second-group streptomycetes. The data obtained confirm the genetic relatedness of the S. cyaneus cluster members and suggest that the structure of the cell wall teichoic acids may serve as one of the taxonomic criteria of the species-level status of streptomycetes.     

Cell wall teichoic acids from Streptomyces daghestanicus VKM Ac-1722T and Streptomyces murinus INA-00524T

The structure of cell wall teichoic acids was studied by chemical methods and NMR spectroscopy in the type strains of two actinomycete species of the "Streptomyces griseoviridis" phenetic cluster: Streptomyces daghestanicus and Streptomyces murinus. S. daghestanicus VKM Ac-1722T contained two polymers having a 1,5-poly(ribitol phosphate) structure. In one of them, the ribitol units had alpha-rhamnopyranose and 3-O-methyl-alpha-rhamnopyranose substituents; in the other, each ribitol unit was carrying 2,4-ketal-bound pyruvic acid. Such polymers were earlier found in the cell walls of Streptomyces roseolus and Nocardiopsis albus, respectively; however, their simultaneous presence in the cell wall has never been reported. The cell wall teichoic acid of Streptomyces murinus INA-00524T was is a 1,5-poly(glucosylpolyol phosphate), whose repeating unit was [-6)-beta-D-glucopyranosyl-(1 --> 2)-glycerol phosphate-(3-P-]. Such a teichoic acid was earlier found in Spirilliplanes yamanashiensis. The 13C NMR spectrum of this polymer is presented for the first time. The results of the present investigation, together with earlier published data, show that the type strains of four species of the "Streptomyces griseoviridis" phenetic cluster differ in the composition and structure of their teichoic acids; thus, teichoic acids may serve as chemotaxonomic markers of the species.     

Teichoic acids from the cell wall of Nocardiopsis prasina BKM Ac-1880T

The cell wall of Nocardiopsis prasina VKM Ac-1880T was found to contain two structurally different teichoic acids: unsubstituted 3,5-poly(ribitol phosphate) and 1,3-poly(glycerol phosphate), substituted at position 2 by 10% with alpha-N-acetylglucosamine and by 5% with O-acetyl groups. The structure of the polymers was studied by chemical analysis and NMR spectroscopy. The results obtained correlate well with 16S rRNA sequence data and confirm the species-specificity of teichoic acids in the genus Nocardiopsis.     

The carbohydrate-containing cell wall polymers of some species from the cluster "Streptomyces lavendulae"

The type strains of the cluster "Streptomyces lavendulae" species with a low level of DNA-DNA relatedness were found to contain different cell-wall carbohydrate polymers, whereas the species of this cluster with a level of DNA-DNA relatedness of about 60% contain similar or identical carbohydrate polymers. The type strains Streptomyces katrae VKM Ac-1220T and S. polychromogenes VKM Ac-1207T synthesize mannan with different amounts of alpha-1,2- and alpha-1,3-substituted mannopyranose units and a small number of 1,3-poly(glycerophosphate) chains. The cell walls of S. lavendulocolor VKM Ac-215T and Streptomyces sp. VKM Ac-2117 were found to contain a hitherto unknown teichuronic acid, whose repeating unit is a disaccharide consisting of diaminomannuronic acid and N-acetylgalactosamine:-->4)-beta-D-ManpNAc3NAcA-(1-->3)-alpha-D- GalpNAc-(1-->. In addition, the cell walls of these two streptomycetes contain beta-glucosylated 1,5-poly(ribitol phosphate). The cell walls of S. virginiae VKM Ac-1218T and S. flavotricini VKM Ac-1277T contain the same poly(glucosyl-glycerophosphate). The results presented in this paper are in accordance with the DNA-DNA relatedness data published earlier and indicate a taxonomic significance of the structure of the cell-wall polysaccharides for the delineation of pheneticall/related Streptomyces species.     

Structure of anionic carbohydrate-containing cell wall polymers in several representatives of the order actinomycetales

A new teichoic acid was identified in the cell walls of Streptomyces griseoviridis VKM Ac-622T, Streptomyces sp. VKM Ac-2091, and Actinoplanes campanulata VKM Ac-1319T. The polymer is poly(glycosylglycerol phosphate). The repeating units of the polymer, alpha-galactopyranosyl-(1-->3)-2-acetamido-2-deoxy-beta-galactopyran+ ++ osyl-(1-->1)-glycerols, are in phosphodiester linkage at C-3 of glycerol and C-6 of galactose. The structures of cell wall teichoic acids in the strains Streptomyces chryseus VKM Ac-200T and "Streptomyces subflavus" VKM Ac-484 similar in morphology and growth characteristics are also identical: 1,5-poly(ribitol phosphate) substituted at C-4(2) by 2-acetamido-2-deoxy-beta-glucopyranosyl residues and 1,3-poly(glycerol phosphate). The taxonomic aspects of these results are discussed.     

Cell wall teichoic acids ofNocardiopsis prasina VKM Ac-1880T

The cell wall ofNocardiopsis prasina VKM Ac-1880^T was found to contain two structurally different teichoic acids: unsubstituted 3,5-poly(ribitol phosphate) and l,3-poly(glycerol phosphate) substituted at position 2 by 10% with α-N-acetylglucosamine and by 5% withO-acetyl groups. The structure of the polymers was studied by chemical analysis and NMR spectroscopy. The results obtained correlate wellwith 16S rRNA sequence data and confirm the species-specificity of teichoic acids in the genusNocardiopsis.     

Cell wall teichoic acids from Streptomyces daghestanicus VKM Ac-1722T and streptomyces murinus INA-00524T

The structure of cell wall teichoic acids was studied by chemical methods and NMR spectroscopy in the type strains of two actinomycete species of the Streptomyces griseoviridis phenetic cluster: streptomyces daghestanicus and streptomyces murinus. S. daghestanicus VKM Ac-1722^t contained two polymers having a 1,5-poly(ribitol phosphate) structure. In one of them, the ribitol units had -rhamnopyranose and 3-O-methyl--rhamnopyranose substituents; in the other, each ribitol unit was carrying 2,4-ketal-bound pyruvic acid. Such polymers were earlier found in the cell walls of Streptomyces roseolus and Nocardiopsis albus, respectively; however, their simultaneous presence in the cell wall has never been reported. The cell wall teichoic acid of Streptomyces murinus INA-00524^T was a 1,5-poly(glucosylpolyol phosphate), whose repeating unit was [-6)--D-glucopyranosyl-(12)-glycerol phosphate-(3-P-]. Such a teichoic acid was earlier found in Spirilliplanes yamanashiensis. The ¹³C NMR spectrum of this polymer is presented for the first time. The results of the present investigation, together with earlier published data, show that the type strains of four species of the Streptomyces griseoviridis phenetic cluster differ in the composition and structure of their teichoic acids; thus, teichoic acids may serve as chemotaxonomic markers of the species.Translated from Mikrobiologiya, Vol. 74, No. 1, 2005, pp. 48–54.Original Russian Text Copyright © 2005 by Streshinskaya, Kozlova, Alferova, Shashkov, Evtushenko.     

A novel type of teichoic acid from the cell wall of Bacillus subtilis VKM B-762

The cell wall of Bacillus subtilis VKM B-762 contains, along with 1,5-poly[4-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)ribitol phosphate], a novel type of glycopolymer involving three types of inter-monomeric bonds in the repeating unit, viz., amide, glycosidic and phosphodiester:Such a structural pattern of natural glycopolymers has been hitherto unknown. This polymer represents a novel type of teichoic acids.     

The occurrence of teichoic acids in streptomycetes

The presence of teichoic acids in a number of streptomycetes led to the conclusion that these biopolymers were widely spread in genus Streptomyces. The nature of the teichoic acid present in the mycelium was determined by extracting it with 10% trichloroacetic acid, precipitating it with ethanol and identifying the precipitated polymer by partial acid and alkali hydrolysis to alditol, alditol phosphates and glycosylalditol phosphates. Most strains examined in this survey contained glycerol or ribitol teichoic acids; in some cases neither type was detected.Structurally teichoic acids closely resemble those of other genera of gram-positive bacteria and in many cases represent poly(glycerol phosphate) and poly(ribitol phosphate) chains. The proportion of alditol residues bearing sugar substituents varied widely.Three species of genus Streptoverticillium contained glycerol teichoic acids. It is belived that some of the data presented in this paper might be used with some success in taxonomic studies of streptomycetes.     

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.     

Late-stage polyribitol phosphate wall teichoic acid biosynthesis in Staphylococcus aureus

Wall teichoic acids are cell wall polymers that maintain the integrity of the cellular envelope and contribute to the virulence of Staphylococcus aureus. Despite the central role of wall teichoic acid in S. aureus virulence, details concerning the biosynthetic pathway of the predominant wall teichoic acid polymer are lacking, and workers have relied on a presumed similarity to the putative polyribitol phosphate wall teichoic acid pathway in Bacillus subtilis. Using high-resolution polyacrylamide gel electrophoresis for analysis of wall teichoic acid extracted from gene deletion mutants, a revised assembly pathway for the late-stage ribitol phosphate-utilizing enzymes is proposed. Complementation studies show that a putative ribitol phosphate polymerase, TarL, catalyzes both the addition of the priming ribitol phosphate onto the linkage unit and the subsequent polymerization of the polyribitol chain. It is known that the putative ribitol primase, TarK, is also a bifunctional enzyme that catalyzes both ribitol phosphate priming and polymerization. TarK directs the synthesis of a second, electrophoretically distinct polyribitol-containing teichoic acid that we designate K-WTA. The biosynthesis of K-WTA in S. aureus strain NCTC8325 is repressed by the accessory gene regulator (agr) system. The demonstration of regulated wall teichoic acid biosynthesis has implications for cell envelope remodeling in relation to S. aureus adhesion and pathogenesis.     

Cell wall teichoic acids of streptomycetes of the phenetic cluster 'Streptomyces fulvissimus'

Structures of the anionic polymers of streptomycetes Streptomyces fulvissimus VKM Ac-994(T), Streptomyces longispororuber VKM Ac-1735(T), Streptomyces aureoveticillatus VKM Ac-48(T) and Streptomyces spectabilis INA 00606 belonging to the phenetic cluster 'S. fulvissimus' were investigated by chemical and NMR spectroscopic methods. A teichoic acid from the cell wall of S. spectabilis INA 00606 was studied in more detail, and this was shown to represent 1,3-poly(glycerol phosphate) substituted with glucosamine (alpha-D-GlcNAc) and L-glutamic acid (non-stoichiometric substitution). For the first time, glutamic acid is identified as an acyl substituent in teichoic acids of streptomycetes. The polymer chain is built of the following fragments: Cell walls of other streptomycetes of the phenocluster under study contain 1,3-poly(glycerol phosphates) with glucosamine as a glycosyl substituent at O-2 of the glycerol phosphate units and L-glutamic acid and lysine as O-2 acyl substituents. Not all amino sugar residues in the polymers of these strains are N-acetylated, and the content of the glucosamine and lysine residues in the polymers of different strains is not the same. Despite certain quantitative differences in the structures of the polymers, one may consider streptomycetes of the phenocluster 'S. fulvissimus' as closely related microorganisms, the details of the structures serving as additional criteria for the determination of the species status of a strain under study.     

Anionic carbohydrate-containing cell wall polymers of Streptomyces melanosporofaciens and related species

The structures of cell wall anionic carbohydrate-containing polymers in Streptomyces melanosporofaciens VKM Ac-1864T and phylogenetically close organisms-S. hygroscopicus subsp. hygroscopicus VKM Ac-831T, S. violaceusniger VKM Ac-583T, S. endus VKM Ac-1331, S. endus VKM Ac-129, and S. rutgersensis subsp. castelarensis VKM Ac-832T--have been comparatively studied by chemical and NMR spectroscopic methods. The natural polymer of a new, previously unknown structure, Kdn (3-deoxy-D-glycero-Dgalacto-non-2-ulopyranosonic acid) with beta-galactose residues at C-9, has been found in the cell walls of all the strains under study. The cell walls of all the studied organisms contain three teichoic acids (TA): a predominant TA (1,3-poly(glycerol phosphate) with N-acetylated alpha-glucosaminyl substitutes by C-2 of glycerol, and minor TAs, 1,3- and 2,3-poly(glycerol phosphate) polymers without substitution. Their chains have O-acetyl and O-lysyl groups. Microorganisms of the above-mentioned species differ in the number of alpha-glucosaminyl substitutes and in the degree of their acetylation in the predominant teichoic acid.     

A polymer with a backbone of 3-deoxy-D-glycero-D-galacto-non-2-ulopyranosonic acid, a teichuronic acid, and a beta-glucosylated ribitol teichoic acid in the cell wall of plant pathogenic Streptomyces sp. VKM Ac-2124.

Structures of cell wall anionic polymers of the strain Streptomyces sp. VKM Ac-2124, a causative agent of potato scab, which is phylogenetically the closest to plant pathogenic species S. setonii and S. caviscabies, were studied. The strain contains three anionic glycopolymers, viz., a teichuronic acid with a disaccharide repeating unit -->6)-alpha-d-Glcp-(1-->4)-beta-d-ManpNAc3NAcA-(1-->, a beta-glucosylated polymer of 3-deoxy-d-glycero-d-galacto-non-2-ulopyranosonic acid (Kdn), and a beta-glucosylated 1,5-poly(ribitol phosphate). The strain studied is the second representative of plant pathogenic streptomycetes inducing potato scab disease, the cell wall anionic polymers of which were shown to contain a Kdn-polymer. Presumably, the presence of Kdn-containing structures in the surface regions of pathogens is essential for their efficient attachment to host plant cells.     

Structural elucidation of the extracellular and cell-wall teichoic acids of Staphylococcus aureus MN8m, a biofilm forming strain

Extracellular teichoic acid, an essential constituent of the biofilm produced by Staphylococcus epidermidis strain RP62A, is also an important constituent of the extracellular matrix of another biofilm producing strain, Staphylococcus aureus MN8m. The structure of the extracellular and cell wall teichoic acids of the latter strain was studied by NMR spectroscopy and capillary electrophoresis-mass spectrometry. Both teichoic acids were found to be a mixture of two polymers, a (1-->5)-linked poly(ribitol phosphate), substituted at the 4-position of ribitol residues with beta-GlcNAc, and a (1-->3)-linked poly(glycerol phosphate), partially substituted with the D-Ala at 2-position of glycerol residue. Such mixture is unusual for S. aureus.     

Structure of a teichoic acid from Nocardioides luteus VKM Ac-1246T cell wall

A teichoic acid from the cell walls of Nocardioides luteus VKM Ac-1246T, a validly described species of the Nocardioides genus, is a 1,5-poly(ribitol phosphate) completely substituted at C-4 by alpha-D-galactopyranosyl residues carrying a 4,6-pyruvate ketal group in R-configuration. The structure of the repeating unit of the polymer is as follows: [figure]. The chain consists of approximately 18 repeating units and six beta-D-galactofuranosyl residues linked in the oligomer by 1,6-glycosidic bonds. The oligomer probably terminates the growing end of the teichoic acid. The structure of the polymer was determined by chemical methods and NMR spectroscopy. This teichoic acid has not been described so far.     

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-β-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-β-galactosaminylglycerol phosphate), the N-acetyl-β-galactosamine being acetalated with pyruvic acid at positions 4 and 6. The major TA of N. synnemataformans is a poly(glycerol phosphate-N-acetyl-β-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.     

Structure of the linkage units between ribitol teichoic acids and peptidoglycan.

The structure of the linkage regions between ribitol teichoic acids and peptidoglycan in the cell walls of Staphylococcus aureus H and 209P and Bacillus subtilis W23 and AHU 1390 was studied. Teichoic acid-linked saccharide preparations obtained from the cell walls by heating at pH 2.5 contained mannosamine and glycerol in small amounts. On mild alkali treatment, each teichoic acid-linked saccharide preparation was split into a disaccharide identified as N-acetylmannosaminyl beta(1----4)N-acetylglucosamine and the ribitol teichoic acid moiety that contained glycerol residues. The Smith degradation of reduced samples of the teichoic acid-linked saccharide preparations from S. aureus and B. subtilis gave fragments characterized as 1,2-ethylenediol phosphate-(glycerolphosphate)3-N-acetylmannosaminyl beta(1----4)N- -acetylxylosaminitol and 1,2-ethylenediolphosphate-(glycerol phosphate)2-N-acetylmannosaminyl beta(1----4)N-acetylxylosaminitol, respectively. The binding of the disaccharide unit to peptidoglycan was confirmed by the analysis of linkage-unit-bound glycopeptides obtained from NaIO4 oxidation of teichoic acid-glycopeptide complexes. Mild alkali treatment of the linkage-unit-bound glycopeptides yielded disaccharide-linked glycopeptides, which gave the disaccharide and phosphorylated glycopeptides on mild acid treatment. Thus, it is concluded that the ribitol teichoic acid chains in the cell walls of the strains of S. aureus and B. subtilis are linked to peptidoglycan through linkage units, (glycerol phosphate)3-N-acetylmannosaminyl beta(1----4)N-acetylglucosamine and (glycerol phosphate)2-N-acetylmannosaminyl beta(1----4)N-acetylglucosamine, respectively.     

Anionic polymers of the cell wall of Streptomyces sp. VKM An-2534

The cell wall of Streptomyces sp. VKM An-2534, the causative agent of common scab in potato tubers, which does not synthesize thaxtomin and is phylogenetically close to phytopathogen Streptomyces setonii sp. ATCC 25497, contains two anionic carbohydrate-containing polymers. The major polymer is teichuronic acid, whose repeating unit is disaccharide --> 4)-beta-D-ManpNAc3NAcyA-(1 --> 3)-alpha-D-GalpNAc-(1-->, where Acy is a residue of acetic or L-glutamic acid. The polymer of such structure has been found in Gram-positive bacteria for the first time. The minor polymer is teichoic acid [1,5-poly(ribitol phosphate)], in which a part of the ribitol residues are glycosylated at C4 with beta-D-Glcp and, probably, with beta-D-GlcpNAc and some residues are O-acylated with Lys residues. The structures were proved by chemical and NMR spectroscopic methods. It is likely that the presence of acidic polysaccharides on the surface of the phytopathogenic streptomycete is necessary for its attachment to the host plant.     

Anionic carbohydrate-containing cell wall polymers of <Emphasis Type="Italic">Streptomyces melanosporofaciens</Emphasis> and related species

The structures of cell wall anionic carbohydrate-containing polymers in Streptomyces melanosporofaciens VKM Ac-1864^T and phylogenetically close organisms—S. hygroscopicus subsp. hygroscopicus VKM Ac-831^T, S. violaceusniger VKM Ac-583^T, S. endus VKM Ac-1331^T, S. endus VKM Ac-129, and S. rutgersensis subsp. castelarensis VKM Ac-832^T—have been comparatively studied by chemical and NMR spectroscopic methods. The natural polymer of a new, previously unknown structure, Kdn (3-deoxy-D-glycero-D-galacto-non-2-ulopyranosonic acid) with β-galactose residues at C-9, has been found in the cell walls of all the strains under study. The cell walls of all the studied organisms contain three teichoic acids (TA): a predominant TA (1,3-poly(glycerol phosphate) with N-acetylated α-glucosaminyl substitutes by C-2 of glycerol, and minor TAs, 1,3-and 2,3-poly(glycerol phosphate) polymers without substitution. Their chains have O-acetyl and O-lysyl groups. Microorganisms of the above-mentioned species differ in the number of α-glucosaminyl substitutes and in the degree of their acetylation in the predominant teichoic acid.