Analysis of the lsi region involved in lipooligosaccharide biosynthesis in Neisseria gonorrhoeae.
The genetic locus (lsi-1) responsible for the transformation of the lipooligosaccharide (LOS)-defective Neisseria gonorrhoeae mutant FA5100 to LOS expression was studied by deletion mutagenesis and sequence analysis. An open reading frame that was preceded by a leader sequence containing regions with the potential to form hairpin loops was identified. A perfect sigma 70 promoter consensus sequence was found upstream from this open reading frame. Promoter function was screened for functionality by using lac fusion cassettes and in vitro transcription-translation analysis. A frameshift mutation in the lsi-1 gene was constructed by site-directed mutagenesis and introduced into the chromosome of FA19, the LOS-expressing isogenic parent strain of FA5100. The mutant was characterized by Southern blotting, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and Western blotting (immunoblotting) and found to be phenotypically identical to FA5100.
Petricoin
E F
EF
Department of Microbiology, University of Maryland, College Park 20742.
Danaher
R J
RJ
Stein
D C
DC
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D015139:Blotting, Southern; D015153:Blotting, Western; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D005091:Exons; D005798:Genes, Bacterial; D008070:Lipopolysaccharides / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009344:Neisseria gonorrhoeae / Q000235:genetics* / Q000378:metabolism; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008070:Lipopolysaccharides; C023023:lipid-linked oligosaccharides