Regulation by a novel protein of the bimodal distribution of lipopolysaccharide in the outer membrane of Escherichia coli.
We report on the cloning and characterization of the rfb gene cluster of the O75 lipopolysaccharide from a urinary tract isolate of Escherichia coli. Deletion cloning defined the minimum region of DNA that expressed the O75 antigen in E. coli host strains to be on a 12.4-kb insert. However, the E. coli strain expressing this region did not produce a polymerized O chain as detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining. A slightly larger DNA clone of 13.4 kb produced a polymerized O chain in E. coli S phi 874 but was found to be abnormal in its distribution over the surface membrane. Normal wild-type E. coli, as with Salmonella spp., has a bimodal distribution of the lipopolysaccharide on the surface which is seen as an abundance of long and short O chains attached to the lipid A-core structure. We found in a region adjacent to the cloned rfb region, and on the opposite side from where the putative polymerase (rfc) is encoded, a novel protein of 35.5 kDa expressed from a 1.75-kb DNA fragment. This protein was shown to complement in trans the E. coli strains carrying plasmids that expressed abnormal, unregulated lipopolysaccharides. The expression of these complemented strains was bimodal in distribution. Mutation of the gene encoding this protein destroyed its ability to regulate O-chain distribution. We propose to call this regulator gene rol, for regulator of O length.
Batchelor
R A
RA
Department of Microbiology and Immunology, Baylor College of Medicine, Texas Medical Center, Houston 77030.
Haraguchi
G E
GE
Hull
R A
RA
Hull
S I
SI
D000942:Antigens, Bacterial / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics*; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D005798:Genes, Bacterial; D005809:Genes, Regulator; D005816:Genetic Complementation Test; D008070:Lipopolysaccharides / Q000378:metabolism*; D019081:O Antigens; D015183:Restriction Mapping
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008070:Lipopolysaccharides; D019081:O Antigens