title: Streptococcus agalactiae capsule polymer length and attachment is determined by the proteins CpsABCD.
abstract: The production of capsular polysaccharides (CPS) or secreted exopolysaccharides is ubiquitous in bacteria, and the Wzy pathway constitutes a prototypical mechanism to produce these structures. Despite the differences in polysaccharide composition among species, a group of proteins involved in this pathway is well conserved. Streptococcus agalactiae (group B Streptococcus; GBS) produces a CPS that represents the main virulence factor of the bacterium and is a prime target in current vaccine development. We used this human pathogen to investigate the roles and potential interdependencies of the conserved proteins CpsABCD encoded in the cps operon, by developing knock-out and functional mutant strains. The mutant strains were examined for CPS quantity, size, and attachment to the cell surface as well as CpsD phosphorylation. We observed that CpsB, -C, and -D compose a phosphoregulatory system where the CpsD autokinase phosphorylates its C-terminal tyrosines in a CpsC-dependent manner. These Tyr residues are also the target of the cognate CpsB phosphatase. An interaction between CpsD and CpsC was observed, and the phosphorylation state of CpsD influenced the subsequent action of CpsC. The CpsC extracellular domain appeared necessary for the production of high molecular weight polysaccharides by influencing CpsA-mediated attachment of the CPS to the bacterial cell surface. In conclusion, although having no impact on cps transcription or the synthesis of the basal repeating unit, we suggest that these proteins are fine-tuning the last steps of CPS biosynthesis (i.e. the balance between polymerization and attachment to the cell wall).
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Chiara
authors: C
authors: Toniolo
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Evita
authors: E
authors: Balducci
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Maria Rosaria
authors: MR
authors: Romano
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Daniela
authors: D
authors: Proietti
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Ilaria
authors: I
authors: Ferlenghi
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Guido
authors: G
authors: Grandi
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Francesco
authors: F
authors: Berti
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy.
authors: Immaculada Margarit Y
authors: IM
authors: Ros
authors: From Novartis Vaccines and Diagnostics, Research Center, Via Fiorentina 1, 53100 Siena, Italy robert.janulczyk@novartis.com.
authors: Robert
authors: R
authors: Janulczyk
mesh_terms: D000818:Animals; D016667:Bacterial Capsules / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002473:Cell Wall / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D015151:Immunoblotting; D051379:Mice; D016253:Microscopy, Immunoelectron; D009154:Mutation; D009876:Operon; D010766:Phosphorylation; D011108:Polymers / Q000378:metabolism*; D011135:Polysaccharides, Bacterial / Q000378:metabolism; D011485:Protein Binding; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013290:Streptococcal Infections / Q000382:microbiology; D013292:Streptococcus agalactiae / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism; D037521:Virulence Factors / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D011108:Polymers; D011135:Polysaccharides, Bacterial; D037521:Virulence Factors; D014443:Tyrosine
keywords: Bacterial Protein Kinase; Capsular Polysaccharide; Carbohydrate Biosynthesis; Glycobiology; Gram-positive Bacteria; Mutagenesis; Streptococcus agalactiae