Cloning of a gonococcal DNA sequence that complements the lipooligosaccharide defects of Neisseria gonorrhoeae 1291d and 1291e.
An isogenic set of gonococcal lipooligosaccharide (LOS) mutants derived from pyocin treatment of Neisseria gonorrhoeae 1291 was used to identify cloned gonococcal DNA fragments. A gene bank from N. gonorrhoeae 1291c chromosomal DNA was made in pLEE10, a shuttle vector that replicates in the gonococcus and Escherichia coli. A plasmid (pSG30) that could transform the LOS mutants 1291d and 1291e to reactivity with monoclonal antibody 3F11 and to production of an LOS component with migration identical to that of the parent, 1291, was identified. pSG30 contains a 9-kb EcoRI fragment. Curing studies indicate that pSG30 encodes gene products that affect LOS biosynthesis in trans. Subcloning identified a 2.6-kb HincII fragment (pSG38) that retained the ability to modify the LOS of 1291d and 1291e. The DNA regions involved in modification of 1291d and 1291e were named lsi-4 and lsi-5, respectively. The region of pSG38 that was involved in LOS modification was further localized by the construction of exonuclease III deletion plasmids. Transformation of these constructs identified a 750-bp fragment that retains the ability to modify 1291e and a 540-bp fragment which retains the ability to modify 1291d.
Sandlin
R C
RC
Department of Microbiology, University of Maryland, College Park 20742.
Apicella
M A
MA
Stein
D C
DC
D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D017353:Gene Deletion; D008070:Lipopolysaccharides / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D009344:Neisseria gonorrhoeae / Q000235:genetics*; D010957:Plasmids; D011709:Pyocins / Q000378:metabolism
D004269:DNA, Bacterial; D008070:Lipopolysaccharides; D011709:Pyocins; C023023:lipid-linked oligosaccharides