GNA2132 is a Neisseria meningitidis antigen of unknown function, discovered by reverse vaccinology, which has been shown to induce bactericidal antibodies in animal models. Here we show that this antigen induces protective immunity in humans and it is recognized by sera of patients after meningococcal disease. The protein binds heparin in vitro through an Arg-rich region and this property correlates with increased survival of the unencapsulated bacterium in human serum. Furthermore, two proteases, the meningococcal NalP and human lactoferrin, cleave the protein upstream and downstream from the Arg-rich region, respectively. We conclude that GNA2132 is an important protective antigen of N. meningitidis and we propose to rename it, N eisserial H eparin B inding A ntigen (NHBA).

We report here the first structure of a member of the IgA protease family at 1.75Å resolution. This protease is a founding member of the Type V (autotransporter) secretion system and is considered a virulence determinant among the bacteria expressing the enzyme. The structure of the enzyme fits that of a classical autotransporter in which several unique domains necessary for protein function are appended to a central, 100 Å long β-helical domain. The N-terminal domain of the IgA protease is found to possess a chymotrypsin-like fold. However, this catalytic domain contains a unique loop D that extends over the active site acting as a lid, gating substrate access. The data presented provide a structural basis for the known ability of IgA proteases to only cleave the P/S/T rich hinge peptide unique to IgA1 in the context of the intact fold of the immunoglobulin. Based upon the structural data as well as molecular modeling, a model is presented that suggests the unique, extended loop D in this IgA protease sterically occludes the active site binding cleft in the absence of immunoglobulin binding. Only in the context of binding of the IgA1 immunoglobulin Fc domain in a valley formed between the N-terminal protease domain and another domain appended to the β-helix spine (domain-2) is the lid stabilized in an open conformation. The stabilization of this open conformation through Fc association subsequently allows access of the hinge peptide to the active site resulting in recognition and cleavage of the substrate.

Haemophilus influenzae is a major cause of otitis media and other respiratory tract disease in children. The pathogenesis of disease begins with colonization of the upper respiratory mucosa, a process that involves evasion of local immune mechanisms and adherence to epithelial cells. Several studies have demonstrated that human milk is protective against H. influenzae colonization and disease. In the present study, we examined the effect of human milk on the H. influenzae IgA1 protease and Hap adhesin, two autotransported proteins that are presumed to facilitate colonization. Our results demonstrated that human milk lactoferrin efficiently extracted the IgA1 protease preprotein from the bacterial outer membrane. In addition, lactoferrin specifically degraded the Hap adhesin and abolished Hap-mediated adherence. Extraction of IgA1 protease and degradation of Hap were localized to the N-lobe of the bilobed lactoferrin molecule and were inhibited by serine protease inhibitors, suggesting that the lactoferrin N-lobe may contain serine protease activity. Additional experiments revealed no effect of lactoferrin on the H. influenzae P2, P5, and P6 outer-membrane proteins, which are distinguished from IgA1 protease and Hap by the lack of an N-terminal passenger domain or an extracellular linker region. These results suggest that human milk lactoferrin may attenuate the pathogenic potential of H. influenzae by selectively inactivating IgA1 protease and Hap, thereby interfering with colonization. Future studies should examine the therapeutic potential of lactoferrin, perhaps as a supplement in infant formulas.