Abstract
Neisseria gonorrhoeae (gonococcus [Ng]) is the causative organism of the sexually transmitted disease gonorrhoea, and no effective vaccine exists currently. In this study, the structure, biological properties, and vaccine potential of the Ng-adhesin complex protein (Ng-ACP) are presented. The crystal structure of recombinant Ng-ACP (rNg-ACP) protein was solved at 1.65 Å. Diversity and conservation of Ng-ACP were examined in different Neisseria species and gonococcal isolates (https://pubmlst.org/neisseria/ database) in silico, and protein expression among 50 gonococcal strains in the Centers for Disease Control and Prevention/Food and Drug Administration (CDCP/FDA) AR Isolate Bank was examined by Western blotting. Murine antisera were raised to allele 10 (strain P9-17)-encoded rNg-ACP protein with different adjuvants and examined by enzyme-linked immunosorbent assay (ELISA), Western blotting, and a human serum bactericidal assay. Rabbit antiserum to rNg-ACP was tested for its ability to prevent Ng-ACP from inhibiting human lysozyme activity in vitro. Ng-ACP is structurally homologous to Neisseria meningitidis ACP and MliC/PliC lysozyme inhibitors. Gonococci expressed predominantly allele 10- and allele 6-encoded Ng-ACP (81% and 15% of isolates, respectively). Murine antisera were bactericidal (titers of 64 to 512, P < 0.05) for the homologous P9-17 strain and heterologous (allele 6) FA1090 strain. Rabbit anti-rNg-ACP serum prevented Ng-ACP from inhibiting human lysozyme with ∼100% efficiency. Ng-ACP protein was expressed by all 50 gonococcal isolates examined with minor differences in the relative levels of expression. rNg-ACP is a potential vaccine candidate that induces antibodies that (i) are bactericidal and (ii) prevent the gonococcus from inhibiting the lytic activity of an innate defense molecule.IMPORTANCENeisseria gonorrhoeae (gonococcus [Ng]) is the causative organism of the sexually transmitted disease gonorrhoea, and the organism is listed by the World Health Organization as a high-priority pathogen for research and development of new control measures, including vaccines. In this study, we demonstrated that the N. gonorrhoeae adhesin complex protein (Ng-ACP) was conserved and expressed by 50 gonococcal strains and that recombinant proteins induced antibodies in mice that killed the bacteria in vitro We determined the structure of Ng-ACP by X-ray crystallography and investigated structural conservation with Neisseria meningitidis ACP and MliC/PliC proteins from other bacteria which act as inhibitors of the human innate defense molecule lysozyme. These findings are important and suggest that Ng-ACP could provide a potential dual target for tackling gonococcal infections.
Highlights
Neisseria gonorrhoeae is the causative organism of the sexually transmitted disease gonorrhoea, and no effective vaccine exists currently
ACP proteins expressed by meningococci, gonococci, and commensal Neisseria species all inhibited human C-type lysozyme (HL) in vitro, and bacterial expression conferred tolerance to HL in vivo, despite Nm-ACP not sharing the conserved MliC/PliC sequence motifs required for lysozyme recognition [15]
To our study that examined the immunogenicity of rNm-ACP [14], we expressed recombinant Ng-ACP (rNg-ACP) with the native N terminus containing the protein leader sequence
Summary
Neisseria gonorrhoeae (gonococcus [Ng]) is the causative organism of the sexually transmitted disease gonorrhoea, and no effective vaccine exists currently. We determined the structure of Ng-ACP by X-ray crystallography and investigated structural conservation with Neisseria meningitidis ACP and MliC/PliC proteins from other bacteria which act as inhibitors of the human innate defense molecule lysozyme These findings are important and suggest that Ng-ACP could provide a potential dual target for tackling gonococcal infections. The goal of an effective preventative gonococcal vaccine has been elusive, and the few vaccines that have entered into clinical trials have largely failed [5] These vaccines included killed whole cells [9], a purified single-antigen pilus-based vaccine [10,11,12], and porin B isolated from the gonococcus (but which was contaminated with lipooligosaccharide [LOS], Rmp, and Opa protein) [13]. ACP proteins expressed by meningococci, gonococci, and commensal Neisseria species all inhibited HL in vitro, and bacterial expression conferred tolerance to HL in vivo, despite Nm-ACP not sharing the conserved MliC/PliC sequence motifs required for lysozyme recognition [15]
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