Expression of Campylobacter hyoilei lipo-oligosaccharide (LOS) antigens in Escherichia coli.
Campylobacter spp. are well recognized as primary pathogens in animals and in people. To isolate and define the genetic regions encoding major surface antigens of Campylobacter hyoilei, genomic DNA of the type strain of the species, RMIT-32A, was cloned into a cosmid vector, pLA2917, in Escherichia coli and the resulting genomic library was screened using antiserum raised to the parent C. hyoilei strain. Six cosmid clones were found to express a series of immunoreactive bands in the 15-25 kDa range. These bands were proteinase K-resistant and were found in the LPS fraction of the cells, suggesting that the recombinant cosmids expressed C. hyoilei lipo-oligosaccharide (LOS) antigen(s). The minimum DNA insert size required for expression of C. hyoilei LOS antigen(s) in E. coli was 11.8 kb. This region was subcloned into the plasmid vector pBR322. The partial sequencing of the 11.8 kb region showed that it contains two ORFs, designated rfbF and rfbP, showing homology with the rfbF gene from Serratia marcescens and the rfbP gene from Salmonella typhimurium. Both genes are involved in LPS synthesis. The region also contained a sequence homologous to the rfaC gene of E. coli and Sal. typhimurium which is involved in core oligosaccharide synthesis.
Korolik
Victoria
V
Department of Applied Biology and Biotechnology, RMIT,GPO Box 2476V, Melbourne 3001,Australia.
Fry
Ben N
BN
Department of Bacteriology, Institute of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Universiteit Utrecht,Yalelaan 1 3584 CL Utrecht,The Netherlands.
Alderton
Malcolm R
MR
Department of Applied Biology and Biotechnology, RMIT,GPO Box 2476V, Melbourne 3001,Australia.
van der Zeijst
Bernard A M
BAM
Department of Bacteriology, Institute of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Universiteit Utrecht,Yalelaan 1 3584 CL Utrecht,The Netherlands.
Coloe
Peter J
PJ
Department of Applied Biology and Biotechnology, RMIT,GPO Box 2476V, Melbourne 3001,Australia.
D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002167:Campylobacter / Q000235:genetics* / Q000276:immunology*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry; D019286:Endopeptidase K; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D016695:Glycosyltransferases / Q000235:genetics; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames / Q000235:genetics; D011993:Recombinant Fusion Proteins; D015183:Restriction Mapping; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008070:Lipopolysaccharides; D011993:Recombinant Fusion Proteins; C110120:RfbF protein, Campylobacter coli; C023023:lipid-linked oligosaccharides; D016695:Glycosyltransferases; C080934:lipooligosaccharide 1,5-heptosyltransferase; D017855:Transferases (Other Substituted Phosphate Groups); C088138:undecaprenyl-phosphate galactosephosphotransferase; D019286:Endopeptidase K