Structural variations and roles of rhamnose-rich cell wall polysaccharides in Gram-positive bacteria
2022; Elsevier BV; Volume: 298; Issue: 10 Linguagem: Inglês
10.1016/j.jbc.2022.102488
ISSN1083-351X
AutoresH. Guérin, Saulius Kulakauskas, Marie‐Pierre Chapot‐Chartier,
Tópico(s)Carbohydrate Chemistry and Synthesis
ResumoRhamnose-rich cell wall polysaccharides (Rha-CWPSs) have emerged as crucial cell wall components of numerous Gram-positive, ovoid-shaped bacteria—including streptococci, enterococci, and lactococci—of which many are of clinical or biotechnological importance. Rha-CWPS are composed of a conserved polyrhamnose backbone with side-chain substituents of variable size and structure. Because these substituents contain phosphate groups, Rha-CWPS can also be classified as polyanionic glycopolymers, similar to wall teichoic acids, of which they appear to be functional homologs. Recent advances have highlighted the critical role of these side-chain substituents in bacterial cell growth and division, as well as in specific interactions between bacteria and infecting bacteriophages or eukaryotic hosts. Here, we review the current state of knowledge on the structure and biosynthesis of Rha-CWPS in several ovoid-shaped bacterial species. We emphasize the role played by multicomponent transmembrane glycosylation systems in the addition of side-chain substituents of various sizes as extracytoplasmic modifications of the polyrhamnose backbone. We provide an overview of the contribution of Rha-CWPS to cell wall architecture and biogenesis and discuss current hypotheses regarding their importance in the cell division process. Finally, we sum up the critical roles that Rha-CWPS can play as bacteriophage receptors or in escaping host defenses, roles that are mediated mainly through their side-chain substituents. From an applied perspective, increased knowledge of Rha-CWPS can lead to advancements in strategies for preventing phage infection of lactococci and streptococci in food fermentation and for combating pathogenic streptococci and enterococci. Rhamnose-rich cell wall polysaccharides (Rha-CWPSs) have emerged as crucial cell wall components of numerous Gram-positive, ovoid-shaped bacteria—including streptococci, enterococci, and lactococci—of which many are of clinical or biotechnological importance. Rha-CWPS are composed of a conserved polyrhamnose backbone with side-chain substituents of variable size and structure. Because these substituents contain phosphate groups, Rha-CWPS can also be classified as polyanionic glycopolymers, similar to wall teichoic acids, of which they appear to be functional homologs. Recent advances have highlighted the critical role of these side-chain substituents in bacterial cell growth and division, as well as in specific interactions between bacteria and infecting bacteriophages or eukaryotic hosts. Here, we review the current state of knowledge on the structure and biosynthesis of Rha-CWPS in several ovoid-shaped bacterial species. We emphasize the role played by multicomponent transmembrane glycosylation systems in the addition of side-chain substituents of various sizes as extracytoplasmic modifications of the polyrhamnose backbone. We provide an overview of the contribution of Rha-CWPS to cell wall architecture and biogenesis and discuss current hypotheses regarding their importance in the cell division process. Finally, we sum up the critical roles that Rha-CWPS can play as bacteriophage receptors or in escaping host defenses, roles that are mediated mainly through their side-chain substituents. From an applied perspective, increased knowledge of Rha-CWPS can lead to advancements in strategies for preventing phage infection of lactococci and streptococci in food fermentation and for combating pathogenic streptococci and enterococci. Bacteria living in complex ecological niches interact with a wide range of other organisms, such as other bacterial members of the local microbiota, bacteriophage predators, and mammalian host cells. Many of these relationships are mediated by glycan structures covering the bacterial surface, which can interface with the outside world via glycan–protein or glycan–glycan interactions (1Mostowy R.J. Holt K.E. Diversity-generating machines: genetics of bacterial sugar-coating.Trends Microbiol. 2018; 26: 1008-1021Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar, 2Porter N.T. Martens E.C. The critical roles of polysaccharides in gut microbial ecology and physiology.Annu. Rev. Microbiol. 2017; 71: 349-369Crossref PubMed Scopus (124) Google Scholar). There are several types of glycans, which exhibit a high level of structural diversity. In Gram-negative bacteria, outer membranes are typically rich in lipopolysaccharides, which are formed of a conserved lipid and variable O-antigen polysaccharide chain (3Whitfield C. Williams D.M. Kelly S.D. Lipopolysaccharide O-antigens-bacterial glycans made to measure.J. Biol. Chem. 2020; 295: 10593-10609Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar). In both Gram-positive and Gram-negative bacteria, capsular polysaccharides (CPS) form a thick outer layer around bacterial cells, while exopolysaccharides are released in the surrounding medium (4Whitfield C. Wear S.S. Sande C. Assembly of bacterial capsular polysaccharides and exopolysaccharides.Annu. Rev. Microbiol. 2020; 74: 521-543Crossref PubMed Scopus (73) Google Scholar). Finally, in Gram-positive bacteria, wall teichoic acids (WTAs) or cell wall polysaccharides (CWPSs) are covalently anchored onto peptidoglycan and partly embedded inside the peptidoglycan layer (5Weidenmaier C. Peschel A. Teichoic acids and related cell-wall glycopolymers in Gram-positive physiology and host interactions.Nat. Rev. Microbiol. 2008; 6: 276-287Crossref PubMed Scopus (527) Google Scholar). The protective Gram-positive cell wall comprises a thick peptidoglycan sacculus, which maintains a bacterium's shape and resistance to internal turgor pressure and protects it against external environmental threats (6Martinez B. Rodriguez A. Kulakauskas S. Chapot-Chartier M.P. Cell wall homeostasis in lactic acid bacteria: Threats and defences.FEMS Microbiol. Rev. 2020; 44: 538-564Crossref PubMed Scopus (17) Google Scholar). The peptidoglycan meshwork is decorated with proteins and other glycopolymers. WTA are commonly found in this setting and can represent up to half the mass of the cell wall like in Bacillus subtilis (7Brown S. Santa Maria Jr., J.P. Walker S. Wall teichoic acids of gram-positive bacteria.Annu. Rev. Microbiol. 2013; 67: 313-336Crossref PubMed Scopus (551) Google Scholar). WTA are anionic glycopolymers, of which the best-characterized examples are made of alditol-phosphate monomers (glycerol- or ribitol-phosphate), possibly including other monosaccharides in their backbone chain and side-chain substituents (mainly sugars and D-alanine). A wide range of functions has been attributed to WTA, including regulation of cell wall growth and morphology, biofilm formation, interactions with bacteriophages, resistance to host cationic antimicrobial peptides, and pathogenicity (7Brown S. Santa Maria Jr., J.P. Walker S. Wall teichoic acids of gram-positive bacteria.Annu. Rev. Microbiol. 2013; 67: 313-336Crossref PubMed Scopus (551) Google Scholar, 8Sumrall E.T. Keller A.P. Shen Y. Loessner M.J. Structure and function of Listeria teichoic acids and their implications.Mol. Microbiol. 2020; 113: 627-637Crossref PubMed Scopus (0) Google Scholar, 9van Dalen R. Peschel A. van Sorge N.M. Wall teichoic acid in Staphylococcus aureus host interaction.Trends Microbiol. 2020; 28: 985-998Abstract Full Text Full Text PDF PubMed Scopus (41) Google Scholar). However, certain Gram-positive bacteria, mainly ovoid-shaped cocci belonging to the genera Streptococcus, Enterococcus, or Lactococcus, synthesize WTA in small amounts or not at all, and most often lack WTA biosynthesis genes. Strikingly, these bacteria instead produce a large amount of CWPS (up to 50% of cell wall mass) that are characterized by a high content of L-rhamnose (Rha), a 6-deoxyhexose commonly found in bacteria but not in humans (10Mistou M.Y. Sutcliffe I.C. van Sorge N.M. Bacterial glycobiology: Rhamnose-containing cell wall polysaccharides in gram-positive bacteria.FEMS Microbiol. Rev. 2016; 40: 464-479Crossref PubMed Scopus (101) Google Scholar). Different appellations are encountered in the literature for these glycopolymers, such as rhamnopolysaccharide, rhamnose polysaccharide, or, when Glc is present in the backbone chain, rhamnose-glucose polysaccharide (RGP). In this review, we will refer to them as rhamnose-rich CWPS (Rha-CWPS). Rha-CWPS were initially studied as group-specific polysaccharides (or C-substances) that enabled the serotype classification of streptococci (groups A to G, further extended to V) referred to as Lancefield classification (11Lancefield R.C. A serological differentiation of human and other groups of hemolytic streptococci.J. Exp. Med. 1933; 57: 571-595Crossref PubMed Scopus (0) Google Scholar). These groups encompass pathogenic species responsible for mild or severe infections in human and animal as well as nonpathogenic species that are part of the commensal human microbiota or widely used in food fermentation. Rha-CWPS share Rha as their major monosaccharide component but differ in their global composition (12Pritchard D.G. Coligan J.E. Speed S.E. Gray B.M. Carbohydrate fingerprints of streptococcal cells.J. Clin. Microbiol. 1981; 13: 89-92Crossref PubMed Scopus (23) Google Scholar). They were found to have a common polyrhamnose core in group A Streptococcus (GAS) (Streptococcus pyogenes), group C Streptococcus (GCS) (including certain strains of Streptococcus dysgalactiae), and Streptococcus mutans, with variation in monosaccharide or disaccharide substituents among species (Fig. 1) (10Mistou M.Y. Sutcliffe I.C. van Sorge N.M. Bacterial glycobiology: Rhamnose-containing cell wall polysaccharides in gram-positive bacteria.FEMS Microbiol. Rev. 2016; 40: 464-479Crossref PubMed Scopus (101) Google Scholar). In group B (Streptococcus agalactiae), a more complex Rha-CWPS, with multiantenna branching structures, was characterized (13Michon F. Brisson J.R. Dell A. Kasper D.L. Jennings H.J. Multiantennary group-specific polysaccharide of group B Streptococcus.Biochemistry. 1988; 27: 5341-5351Crossref PubMed Scopus (44) Google Scholar). Because of these affiliations, these Rha-CWPS are also called group A carbohydrates (GAC), group B carbohydrates (GBC), or group C carbohydrates (GCC). It is important to note that the Lancefield serological classification is not able to distinguish streptococci at the species level as defined nowadays on the basis of genomic sequences; this means that strains of different genetic species may synthesize identical Rha-CWPS, whereas within other species, different strains express different types of Rha-CWPS. In more recent years, Rha-CWPS have also been structurally characterized in nonpathogenic food lactic acid bacteria such as Lactococcus lactis (formerly Streptococcus lactis, group N Streptococcus) (14Sadovskaya I. Vinogradov E. Courtin P. Armalyte J. Meyrand M. Giaouris E. et al.Another brick in the wall: A rhamnan polysaccharide trapped inside peptidoglycan of Lactococcus lactis.MBio. 2017; 8: e01303-e01317Crossref PubMed Scopus (33) Google Scholar) and Streptococcus thermophilus (15Lavelle K. Sadovskaya I. Vinogradov E. Kelleher P. Lugli G.A. Ventura M. et al.Brussowvirus SW13 requires a cell surface-associated polysaccharide to recognise its Streptococcus thermophilus host.Appl. Environ. Microbiol. 2021; 88 (e01723-21)PubMed Google Scholar, 16McDonnell B. Hanemaaijer L. Bottacini F. Kelleher P. Lavelle K. Sadovskaya I. et al.A cell wall-associated polysaccharide is required for bacteriophage adsorption to the Streptococcus thermophilus cell surface.Mol. Microbiol. 2020; 114: 31-45Crossref PubMed Scopus (0) Google Scholar). In these bacteria, Rha-CWPS have received significant attention for their role as receptors for numerous infecting bacteriophages that threaten milk fermentation in the food industry (17Mahony J. Cambillau C. van Sinderen D. Host recognition by lactic acid bacterial phages.FEMS Microbiol. Rev. 2017; 41: S16-S26Crossref PubMed Scopus (30) Google Scholar, 18Romero D.A. Magill D. Millen A. Horvath P. Fremaux C. Dairy lactococcal and streptococcal phage-host interactions: An industrial perspective in an evolving phage landscape.FEMS Microbiol. Rev. 2020; 44: 909-932Crossref PubMed Scopus (10) Google Scholar, 19Lavelle K. Sinderen D.V. Mahony J. Cell wall polysaccharides of Gram positive ovococcoid bacteria and their role as bacteriophage receptors.Comput. Struct. Biotechnol. J. 2021; 19: 4018-4031Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar). Finally, in pathogenic Enterococcus faecalis (formerly Streptococcus faecalis, group D Streptococcus), Rha-CWPS (also named enterococcal polysaccharide antigen, EPA) was found to influence virulence and colonizing potential in the host gut (20Rigottier-Gois L. Madec C. Navickas A. Matos R.C. Akary-Lepage E. Mistou M.Y. et al.The surface rhamnopolysaccharide epa of Enterococcus faecalis is a key determinant of intestinal colonization.J. Infect Dis. 2015; 211: 62-71Crossref PubMed Scopus (49) Google Scholar, 21Teng F. Singh K.V. Bourgogne A. Zeng J. Murray B.E. Further characterization of the epa gene cluster and Epa polysaccharides of.Enterococcus Faecalis. Infect Immun. 2009; 77: 3759-3767Crossref PubMed Scopus (0) Google Scholar, 22Ramos Y. Sansone S. Morales D.K. Sugarcoating it: enterococcal polysaccharides as key modulators of host-pathogen interactions.PLoS Pathog. 2021; 17e1009822Crossref Scopus (2) Google Scholar). In all three of these latter species, Rha-CWPS are complex heteropolysaccharides made of a polyrhamnose chain substituted with variable oligosaccharide or polysaccharide chains (Fig. 1). Rha-CWPS are not present in all streptococci. For example, they are absent in Streptococcus pneumoniae, whose cell wall instead contains WTA and lipoteichoic acids (LTAs) (23Vollmer W. Massidda O. Tomasz A. The cell wall of Streptococcus pneumoniae.Microbiol. Spectr. 2019; 7 (GPP3-0018-2018)Crossref PubMed Scopus (23) Google Scholar) and is covered with a capsule, the structure of which is highly variable among strains and allows the definition of at least 90 serotypes (24Yother J. Capsules of Streptococcus pneumoniae and other bacteria: Paradigms for polysaccharide biosynthesis and regulation.Annu. Rev. Microbiol. 2011; 65: 563-581Crossref PubMed Scopus (216) Google Scholar). However, a Rha-CWPS was also recently characterized in a more distantly related bacterial species, Ruminococcus gnavus, an anaerobic ovoid-shaped Gram-positive bacterium that is a component of the human gut microbiota (25Henke M.T. Kenny D.J. Cassilly C.D. Vlamakis H. Xavier R.J. Clardy J. Ruminococcus gnavus, a member of the human gut microbiome associated with Crohn's disease, produces an inflammatory polysaccharide.Proc. Natl. Acad. Sci. U. S. A. 2019; 116: 12672-12677Crossref PubMed Scopus (267) Google Scholar). This finding suggests that the occurrence of Rha-CWPS could be broader than previously anticipated. The importance and distinctiveness of Rha-CWPS in Gram-positive bacteria were previously presented in a comprehensive review (10Mistou M.Y. Sutcliffe I.C. van Sorge N.M. Bacterial glycobiology: Rhamnose-containing cell wall polysaccharides in gram-positive bacteria.FEMS Microbiol. Rev. 2016; 40: 464-479Crossref PubMed Scopus (101) Google Scholar). Since then, however, significant progress has been achieved. Novel structures have been identified, subtle but essential modifications of known structures have been discovered, and significant advances have been made regarding their biosynthesis and roles in bacterial physiology and interactions with the environment. Here, we present an update on what is currently known regarding the structure, biosynthesis, and roles of Rha-CWPS, focusing on a few well-characterized ovoid-shaped bacterial species. We describe the complex and diversified arsenal of enzymatic activities dedicated to side-chain substituent biosynthesis, including multicomponent transmembrane glycosylation systems. These systems involve GT-C–fold glycosyltransferases using polyprenyl-monophosphate–linked sugar donors. We discuss emerging data that reveal the importance of Rha-CWPS in bacterial cell wall architecture and cell division. Finally, we summarize their roles, with a particular emphasis on the influence of side-chain substituents, as bacteriophage receptors and in host interactions. Among different bacterial species, Rha-CWPS share several common structural features, upon which an astonishing variety of chemical refinements are constructed by sophisticated glycopolymer modification systems. The polyrhamnose core is relatively conserved, while the side-chain substituents are characterized by extensive diversity in both size and structure among and even within species. The constituent repeating units of the rhamnose-rich backbone differ with respect to the number of L-Rha residues (between 2 and 5) and the glycosidic linkages between them (Fig. 1A). The most conserved subunit structure is made of two L-Rha residues with alternating α-1,2- and α-1-3-L-Rha (Fig. 2). In certain cases, the repeating unit contains one glucose (Glc), thus accounting for the name RGP. For the sake of simplicity in this text, we will refer to this linear Rha-rich backbone as rhamnan in all cases. The rhamnan polymer is synthesized with a linker unit made of GlcNAc, which may be N-deacetylated to glucosamine (GlcNH2) in certain cases (26Rush J.S. Parajuli P. Ruda A. Li J. Pohane A.A. Zamakhaeva S. et al.PplD is a de-N-acetylase of the cell wall linkage unit of streptococcal rhamnopolysaccharides.Nat. Commun. 2022; 13: 590Crossref PubMed Scopus (0) Google Scholar). In addition, a cap structure made of sugar has been suggested by experimental data (14Sadovskaya I. Vinogradov E. Courtin P. Armalyte J. Meyrand M. Giaouris E. et al.Another brick in the wall: A rhamnan polysaccharide trapped inside peptidoglycan of Lactococcus lactis.MBio. 2017; 8: e01303-e01317Crossref PubMed Scopus (33) Google Scholar, 27Vinogradov E. Sadovskaya I. Courtin P. Kulakauskas S. Grard T. Mahony J. et al.Determination of the cell wall polysaccharide and teichoic acid structures from Lactococcus lactis IL1403.Carbohydr. Res. 2018; 462: 39-44Crossref PubMed Scopus (17) Google Scholar) but it is not known at present whether this is a conserved feature of Rha-CWPS. Two classes of Rha-CWPS can be distinguished based on their side-chain substituents: the first group has only simple monosaccharide or disaccharide substituents, while the second possesses complex oligosaccharide or polysaccharide chain substituents (Fig. 1B). In S. pyogenes (GAS), GlcNAc monosaccharide is grafted onto the polyrhamnose backbone in all serotypes and is the immunodominant epitope, allowing serological discrimination of GAS (28van Sorge N.M. Cole J.N. Kuipers K. Henningham A. Aziz R.K. Kasirer-Friede A. et al.The classical lancefield antigen of group A Streptococcus is a virulence determinant with implications for vaccine design.Cell Host Microbe. 2014; 15: 729-740Abstract Full Text Full Text PDF PubMed Scopus (89) Google Scholar). Interestingly, the GlcNAc substituents were recently found to be themselves (partly) modified by a glycerol-phosphate (GroP) group (29Edgar R.J. van Hensbergen V.P. Ruda A. Turner A.G. Deng P. Le Breton Y. et al.Discovery of glycerol phosphate modification on streptococcal rhamnose polysaccharides.Nat. Chem. Biol. 2019; 15: 463-471Crossref PubMed Scopus (36) Google Scholar). It is likely that this modification had not been detected earlier because of the acidic conditions used for extracting Rha-CWPS from the cell wall, which lead to GroP loss. In S. mutans, Glc is the common substituent in three different serotypes, each with a different chemical bond between Glc and Rha: α-1,2- (serotype c), β-1,2- (serotype e), and α-1,3 (serotype f) (30Nakano K. Ooshima T. Serotype classification of Streptococcus mutans and its detection outside the oral cavity.Future Microbiol. 2009; 4: 891-902Crossref PubMed Scopus (85) Google Scholar). In addition, GroP was recently shown to be attached to the serotype c carbohydrate and possibly to others as well (31Zamakhaeva S. Chaton C.T. Rush J.S. Ajay Castro S. Kenner C.W. Yarawsky A.E. et al.Modification of cell wall polysaccharide guides cell division in.Streptococcus Mutans. Nat. Chem. Biol. 2021; 17: 878-887Crossref PubMed Scopus (8) Google Scholar). In GCS, disaccharide substituents comprise two GalNAc units, with a certain proportion of subunits devoid of substituents (32Neiwert O. Holst O. Duda K.A. Structural investigation of rhamnose-rich polysaccharides from Streptococcus dysgalactiae bovine mastitis isolate.Carbohydr. Res. 2014; 389: 192-195Crossref PubMed Scopus (14) Google Scholar), whereas in group G Streptococcus, they are composed of Rha-GalNAc (33Pritchard D.G. Rener B.P. Furner R.L. Huang D.H. Krishna N.R. Structure of the group G streptococcal polysaccharide.Carbohydr. Res. 1988; 173: 255-262Crossref PubMed Scopus (0) Google Scholar) (Fig. 1B). In the second class of Rha-CWPS, substituents are far more complex. In S. agalactiae (GBS), a multiantenna polysaccharide was structurally characterized several decades ago as GBC common to all GBS strains (13Michon F. Brisson J.R. Dell A. Kasper D.L. Jennings H.J. Multiantennary group-specific polysaccharide of group B Streptococcus.Biochemistry. 1988; 27: 5341-5351Crossref PubMed Scopus (44) Google Scholar). The tri-Rha terminal motifs of the phospho-octasaccharide subunits constitute the major antigenic determinant of GBS serological classification. More recently, two elements were highlighted in this complex structure: a stem oligosaccharide made of five Rha residues and a substituting multiantenna structure made of repeated octasaccharide that contains one phosphate (10Mistou M.Y. Sutcliffe I.C. van Sorge N.M. Bacterial glycobiology: Rhamnose-containing cell wall polysaccharides in gram-positive bacteria.FEMS Microbiol. Rev. 2016; 40: 464-479Crossref PubMed Scopus (101) Google Scholar) (Fig. 1B). The gene cluster involved in GBC biosynthesis is part of the S. agalactiae core genome (34Tettelin H. Masignani V. Cieslewicz M.J. Donati C. Medini D. Ward N.L. et al.Genome analysis of multiple pathogenic isolates of Streptococcus agalactiae: Implications for the microbial "pan-genome".Proc. Natl. Acad. Sci. U. S. A. 2005; 102: 13950-13955Crossref PubMed Scopus (0) Google Scholar), in agreement with the conserved GBC structure among strains. In contrast, different strains of L. lactis synthesize complex Rha-CWPS that exhibit high structural diversity (35Theodorou I. Courtin P. Palussiere S. Kulakauskas S. Bidnenko E. Pechoux C. et al.A dual-chain assembly pathway generates the high structural diversity of cell-wall polysaccharides in.Lactococcus Lactis. J. Biol. Chem. 2019; 294: 17612-17625Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar) (Fig. 1B). This structural diversity parallels the genetic variations found within the large gene cluster responsible for their biosynthesis (36Mahony J. Frantzen C. Vinogradov E. Sadovskaya I. Theodorou I. Kelleher P. et al.The CWPS Rubik's cube: linking diversity of cell wall polysaccharide structures with the encoded biosynthetic machinery of selected Lactococcus lactis strains.Mol. Microbiol. 2020; 114: 582-596Crossref PubMed Scopus (0) Google Scholar). To date, more than 10 chemical structures have been established by 2D-NMR spectroscopy studies in different strains. The variable part of the structures determines their sensitivity to bacteriophages, which suggests the influence of a coevolutionary arms race between phage receptor-binding proteins and their saccharidic ligands at the bacterial surface (17Mahony J. Cambillau C. van Sinderen D. Host recognition by lactic acid bacterial phages.FEMS Microbiol. Rev. 2017; 41: S16-S26Crossref PubMed Scopus (30) Google Scholar, 19Lavelle K. Sinderen D.V. Mahony J. Cell wall polysaccharides of Gram positive ovococcoid bacteria and their role as bacteriophage receptors.Comput. Struct. Biotechnol. J. 2021; 19: 4018-4031Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar). In general, Rha-CWPS in L. lactis appear to be composed of a rhamnan chain (14Sadovskaya I. Vinogradov E. Courtin P. Armalyte J. Meyrand M. Giaouris E. et al.Another brick in the wall: A rhamnan polysaccharide trapped inside peptidoglycan of Lactococcus lactis.MBio. 2017; 8: e01303-e01317Crossref PubMed Scopus (33) Google Scholar) that is covalently linked to a variable oligosaccharide or polysaccharide component exposed at the bacterial surface (Fig. 1B) (37Chapot-Chartier M.P. Vinogradov E. Sadovskaya I. Andre G. Mistou M.Y. Trieu-Cuot P. et al.The cell surface of Lactococcus lactis is covered by a protective polysaccharide pellicle.J. Biol. Chem. 2010; 285: 10464-10471Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar, 38Ainsworth S. Sadovskaya I. Vinogradov E. Courtin P. Guerardel Y. Mahony J. et al.Differences in lactococcal cell wall polysaccharide structure are major determining factors in bacteriophage sensitivity.MBio. 2014; 5 (e00880-14)Crossref PubMed Scopus (75) Google Scholar). In several strains, the rhamnan chain is made of identical tri-Rha subunits, while in L. lactis IL1403, it is a disaccharide subunit (27Vinogradov E. Sadovskaya I. Courtin P. Kulakauskas S. Grard T. Mahony J. et al.Determination of the cell wall polysaccharide and teichoic acid structures from Lactococcus lactis IL1403.Carbohydr. Res. 2018; 462: 39-44Crossref PubMed Scopus (17) Google Scholar) (Fig. 2) and, in strain UC509.9, a tetrasaccharide with three Rha and one Glc (39Vinogradov E. Sadovskaya I. Grard T. Murphy J. Mahony J. Chapot-Chartier M.P. et al.Structural studies of the cell wall polysaccharide from Lactococcus lactis UC509.9.Carbohydr. Res. 2018; 461: 25-31Crossref PubMed Scopus (14) Google Scholar) (Fig. 1A). The rhamnan chain may also be substituted with lateral monosaccharide Glc; this was found in very low amounts in strains MG1363 and 3107 (40Theodorou I. Courtin P. Sadovskaya I. Palussiere S. Fenaille F. Mahony J. et al.Three distinct glycosylation pathways are involved in the decoration of Lactococcus lactis cell wall glycopolymers.J. Biol. Chem. 2020; 295: 5519-5532Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar) but was present on each tri-Rha subunit in strain SK11 (Fig. 1) (36Mahony J. Frantzen C. Vinogradov E. Sadovskaya I. Theodorou I. Kelleher P. et al.The CWPS Rubik's cube: linking diversity of cell wall polysaccharide structures with the encoded biosynthetic machinery of selected Lactococcus lactis strains.Mol. Microbiol. 2020; 114: 582-596Crossref PubMed Scopus (0) Google Scholar). In most described structures, the polysaccharide substituents are made of pentasaccharide or hexasaccharide subunits containing a phosphate group (Fig. 1B) (37Chapot-Chartier M.P. Vinogradov E. Sadovskaya I. Andre G. Mistou M.Y. Trieu-Cuot P. et al.The cell surface of Lactococcus lactis is covered by a protective polysaccharide pellicle.J. Biol. Chem. 2010; 285: 10464-10471Abstract Full Text Full Text PDF PubMed Scopus (132) Google Scholar, 38Ainsworth S. Sadovskaya I. Vinogradov E. Courtin P. Guerardel Y. Mahony J. et al.Differences in lactococcal cell wall polysaccharide structure are major determining factors in bacteriophage sensitivity.MBio. 2014; 5 (e00880-14)Crossref PubMed Scopus (75) Google Scholar, 41Farenc C. Spinelli S. Vinogradov E. Tremblay D. Blangy S. Sadovskaya I. et al.Molecular insights on the recognition of a Lactococcus lactis cell wall pellicle by the phage 1358 receptor binding protein.J. Virol. 2014; 88: 7005-7015Crossref PubMed Scopus (49) Google Scholar). The oligosaccharide substituents consist of a trisaccharide in strain IL1403 (27Vinogradov E. Sadovskaya I. Courtin P. Kulakauskas S. Grard T. Mahony J. et al.Determination of the cell wall polysaccharide and teichoic acid structures from Lactococcus lactis IL1403.Carbohydr. Res. 2018; 462: 39-44Crossref PubMed Scopus (17) Google Scholar) or a hexasaccharide in strain UC509.9 (39Vinogradov E. Sadovskaya I. Grard T. Murphy J. Mahony J. Chapot-Chartier M.P. et al.Structural studies of the cell wall polysaccharide from Lactococcus lactis UC509.9.Carbohydr. Res. 2018; 461: 25-31Crossref PubMed Scopus (14) Google Scholar), with notable nonstoichiometric GroP substituents in both strains. In S. thermophilus, the structures of two Rha-CWPS (called RGP in the original studies) established by NMR revealed an identical backbone structure, with subunits made of two L-Rha and one Glc, together with lateral oligosaccharide side chains containing similar sugars but different structures (15Lavelle K. Sadovskaya I. Vinogradov E. Kelleher P. Lugli G.A. Ventura M. et al.Brussowvirus SW13 requires a cell surface-associated polysaccharide to recognise its Streptococcus thermophilus host.Appl. Environ. Microbiol. 2021; 88 (e01723-21)PubMed Google Scholar, 16McDonnell B. Hanemaaijer L. Bottacini F. Kelleher P. Lavelle K. Sadovskaya I. et al.A cell wall-associated polysaccharide is required for bacteriophage adsorption to the Streptococcus thermophilus cell surface.Mol. Microbiol. 2020; 114: 31-45Crossref PubMed Scopus (0) Google Scholar) (Fig. 1). Finally, in strain V583 of E. faecalis, a complex structure was described for EPA (another type of Rha-CWPS), with a rhamnan backbone covalently bound to teichoic acid–like chains (42Guerardel Y. Sadovskaya I. Maes E. Furlan S. Chapot-Chartier M.P. Mesnage S. et al.Complete structure of the enterococcal polysaccharide antigen (EPA) of vancomycin-resistant Enterococcus faecalis V583 reveals that EPA decorations are teichoic acids covalently linked to a rhamnopolysaccharide backbone.mBio. 2020; 11 (e00277-20)Crossref Scopus (18) Google Scholar) (Fig. 1). Since the epa chromosomal locus exhibits variability among strains, structural diversity is likely in the dec
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