Immunoproteomic identification of the hypothetical protein NMB1468 as a novel lipoprotein ubiquitous in Neisseria meningitidis with vaccine potential

Abstract

Many potential vaccine candidates for serogroup B Neisseria meningitidis (NMB) have been identified by reverse vaccinology, a genome-based approach. However, some candidates may be unseen owing to uncertain annotation or their peculiar properties. In this study, we describe the preparation and identification of a novel lipoprotein expressed in all meningococcal strains tested. mAb were first prepared from mice immunized with a meningococcal B strain isolated in Taiwan. Total proteins from the immunizing strain were separated by 2-DE in duplicate. Clone 4-7-3, which crossreacted to 174 tested meningococcal isolates, was used as the primary antibody for Western blotting. The immunoreactive spot was identified by LC-mass spectrometric analysis of the corresponding spot from the silver-stained gel and confirmed by molecular biology approach to be a novel lipoprotein encoded by the hypothetical NMB1468 gene. The potential use of this protein, designated Ag473/NMB1468, as a vaccine component was evaluated using the recombinant protein produced in Escherichia coli. Immunized mice were found to be protected from developing meningococcal disease after intraperitoneal inoculation with a lethal dose of meningococcal strain Nm22209, suggesting that Ag473/NMB1468 may be a promising vaccine candidate. This study also demonstrates the usefulness of the immunoproteomic approach in identification of novel vaccine candidates.

Genetic, Structural, and Antigenic Analyses of Glycan Diversity in the O-Linked Protein Glycosylation Systems of HumanNeisseriaSpecies

“…In this context, it is interesting that other studies have reported evidence indicating that some of the glycoproteins identified in N. gonorrhoeae and now in N. meningitidis are exposed at the bacterial surface. These glycoproteins include a putative peptidyl-prolylisomerase (NGO1225) (38), GNA1946 (45), Ag473 (NGO1043) (28), and the nitrite reductase AniA (NGO1276) (37). It is also worth mentioning that the original evidence that AniA is expressed in vivo was derived from detection of AniA-reactive antibodies in diseased patients but not in normal human sera (12).…”

High-Throughput Identification of New Protective Antigens from a Yersinia pestis Live Vaccine by Enzyme-Linked Immunospot Assay

“…The development of protein microarrays and immunoproteomics let us identify novel immunogens that induce humoral immune responses in a high-throughput manner (1,12,19,21,27,28). The approach to identify T-cell antigens on a large scale is under way (10,14).…”

Vaccinology in the genome era

“…A number of methods have been developed to enable high-throughput display of the proteome of a pathogen to the host immune system. Immunomics (also known as serological proteome analysis [SERPA]) combines proteomic-based approaches with serological analysis and has been widely applied for antigen discovery and vaccine development (72)(73)(74). Applied to S. aureus, this approach led to the identification of 15 highly immunogenic proteins, including known and novel vaccine candidates (75).…”