Selection and immunochemical analysis of lipooligosaccharide mutants of Neisseria gonorrhoeae

Abstract

The identification of enterobacterial mutants that contain alterations in the lipopolysaccharide (LPS) oligosaccharide core structure facilitated the development of the model of the physicochemical and immunochemical structures of enteric LPS. Results of recent immunochemical studies have suggested that the structural model of the lipooligosaccharides (LOSs) of Neisseria gonorrhoeae may differ from the enteric LPS model. The difficulties in the analysis of the wild-type gonococcal LOS have precluded understanding of the precise nature of the LOS structure. This study was undertaken to isolate a series of mutants of N. gonorrhoeae 1291 that had sequential saccharide deletions in the LOS. Results of preliminary studies suggested that the pyocin, designated pyocin C, allowed selection of gonococci with such mutant LOS structures. Results also indicated that the receptor for pyocin C binding was an LOS component. Pyocin C selection led to the isolation of five strains with LOS patterns on sodium dodecyl sulfate-polyacrylamide gels which differed from the LOS of parent strain 1291. In this system, the Mr of the parent LOS was 4,715, while the LOSs from the mutant strains demonstrated progressive saccharide deletions, with Mrs of 4,230, 4,089, 3,627, 3,262, and 3,197. Protein patterns of these mutants on sodium dodecyl sulfate-polyacrylamide gels were qualitatively similar to those of the parent strains. Results of studies with five monoclonal antibodies specific for neisserial LOS indicated that shared as well as unique epitopes were present on the mutant LOSs. Results of ketodeoxyoctonate analysis of the mutant LOSs indicated that the majority of the ketodeoxyoctonate residues may be substituted on C-4 or C-5. Chemical and immunological analysis of such LOS mutants should expedite the development of the model for the structure of gonococcal LOS.

Topographical alterations in proteins I of Neisseria gonorrhoeae correlated with lipooligosaccharide variation

“…The relatively poor silver staining of the LOS of strains WS2 and WS3 was noted previously by Shafer et al (1986) for the LOS of parental strain WS1. N. mutant of FA102 (Shafer e( a/., 1986) FA699XWS1; this study FA899XWS1; this study WS2XFA102; this study WS2XFA102: this study gonorrhoeae lipooligosaccharide migration in SDS-PAGE gels appears to correspond to the Salmonella chemotype mutants Ra (rough) to Re (deep rough) (Dudas and Apicella, 1988), suggesting that WS2 and WS3 have 'deep rough' LOS types wheres WS4 and WS5 have 'rough' LOS types. The LOS profiles of WS4 and WS5 also appeared to differ, perhaps because of the presence of a locus tightly associated with, but distinct from, the locus involved in P.IB9 synthesis (Stephens and Shafer, 1987).…”

“…'Forcorrespondence. (Bergstrom, etal.. 1986;Sparling, etal.. 1986) and proteins II (P.lls) (Sparling, ef a/., 1986) exhibit both phase and antigenic variation within and among strains, while lipooligosaccharide (LOS) can vary considerably within a single strain (Dudas and Apicella, 1988). The major OM protein, protein I (P.I.…”

Infection of human urethral epithelium with Neisseria gonorrhoeae elicits an upregulation of host anti-apoptotic factors and protects cells from staurosporine-induced apoptosis

“…Neisseria gonorrhoeae strain 1291 was used in these studies. This strain was originally isolated from a male with gonococcal urethritis, and has been characterized to express a single species of LOS (Dudas and Apicella, 1988;John et al, 1991). Strain 1291 was grown overnight (37∞C/5% CO 2 ) on GC agar, and P + (pilus-positive), Opa + (opacity-associated protein-positive) gonococci were selected on the basis of colony morphology.…”

Receptor‐mediated endocytosis of Neisseria gonorrhoeae into primary human urethral epithelial cells: the role of the asialoglycoprotein receptor

“…Neisseria gonorrhoeae strain 1291 was used for these studies. This strain expresses a single species of LOS that has been characterized and described previously (Dudas and Apicella, 1988;John et al, 1991). Construction and characterization of N. gonorrhoeae strain 1291-R6, a phosphoglucomutase (pgm) mutant that lacks the lacto-N-neotetraose moiety found on the parent strain LOS, have been described previously (Zhou et al, 1994).…”