title: Involvement of the rml locus in core oligosaccharide and O polysaccharide assembly in Pseudomonas aeruginosa.
abstract: L-Rhamnose (L-Rha) is a component of the lipopolysaccharide (LPS) core, several O antigen polysaccharides, and the cell surface surfactant rhamnolipid of Pseudomonas aeruginosa. In this study, four contiguous genes (rmlBDAC) responsible for the synthesis of dTDP-L-Rha in P. aeruginosa have been cloned and characterized. Non-polar chromosomal rmlC mutants were generated in P. aeruginosa strains PAO1 (serotype O5) and PAK (serotype O6) and LPS extracted from the mutants was analysed by SDS-PAGE and Western immunoblotting. rmlC mutants of both serotype O5 and serotype O6 synthesized a truncated core region which was unable to act as an attachment point for either A-band or B-band O antigen. A rmd rmlC PAO1 double mutant (deficient in biosynthesis of both D-Rha and L-Rha) was constructed to facilitate structural analysis of the mutant core region. This strain has an incomplete core oligosaccharide region and does not produce A-band O antigen. These results provide the genetic and structural evidence that L-Rha is the receptor on the P. aeruginosa LPS core for the attachment of O polysaccharides. This is the first report of a genetically defined mutation that affects the synthesis of a single sugar in the core oligosaccharide region of P. aeruginosa LPS, and provides further insight into the mechanisms of LPS biosynthesis and assembly in this bacterium.
authors.affiliation: Canadian Bacterial Diseases Network, Networks of Centers of Excellence, Heritage Medical Research Building, Hospital Drive, Calgary, Alberta, Canada T2N 4N12.|Department of Microbiology, University of Guelph, Guelph, Ontario, Canada, N1G 2W11.
authors.forename: Rahim
authors.initials: R
authors.lastname: Rahim
authors.affiliation: Center for Infection and Biomaterials Research, NU13-143, Toronto General Hospital, Toronto, Ontario, Canada M5G 2C43.
authors.forename: Lori L
authors.initials: LL
authors.lastname: Burrows
authors.affiliation: Institute for Biological Sciences, National Research Council, Ottawa, Ontario, Canada K1A OR64.|Canadian Bacterial Diseases Network, Networks of Centers of Excellence, Heritage Medical Research Building, Hospital Drive, Calgary, Alberta, Canada T2N 4N12.
authors.forename: Mario A
authors.initials: MA
authors.lastname: Monteiro
authors.affiliation: Institute for Biological Sciences, National Research Council, Ottawa, Ontario, Canada K1A OR64.|Canadian Bacterial Diseases Network, Networks of Centers of Excellence, Heritage Medical Research Building, Hospital Drive, Calgary, Alberta, Canada T2N 4N12.
authors.forename: Malcolm B
authors.initials: MB
authors.lastname: Perry
authors.affiliation: Canadian Bacterial Diseases Network, Networks of Centers of Excellence, Heritage Medical Research Building, Hospital Drive, Calgary, Alberta, Canada T2N 4N12.|Department of Microbiology, University of Guelph, Guelph, Ontario, Canada, N1G 2W11.
authors.forename: Joseph S
authors.initials: JS
authors.lastname: Lam
mesh_terms: D001483:Base Sequence; D002237:Carbohydrate Dehydrogenases / Q000235:genetics / Q000378:metabolism; D002238:Carbohydrate Epimerases / Q000235:genetics / Q000378:metabolism; D002876:Chromosomes, Bacterial / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D005798:Genes, Bacterial; D006836:Hydro-Lyases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism; D019081:O Antigens / Q000096:biosynthesis*; D009844:Oligosaccharides / Q000096:biosynthesis*; D009876:Operon; D010957:Plasmids / Q000235:genetics; D011550:Pseudomonas aeruginosa / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D012210:Rhamnose / Q000378:metabolism; D012703:Serotyping; D001426:Bacterial Proteins
chemical_list: D002237:Carbohydrate Dehydrogenases; D002238:Carbohydrate Epimerases; D017931:DNA Primers; D006836:Hydro-Lyases; D009713:Nucleotidyltransferases; D019081:O Antigens; D009844:Oligosaccharides; D012210:Rhamnose; C107144:dTDP-4-dehydrorhamnose reductase; C020587:glucose-1-phosphate thymidylyltransferase; C022771:dTDPglucose 4,6-dehydratase; C020092:dTDP-4-ketorhamnose 3,5-epimerase; D001426:Bacterial Proteins