Consensus computational network analysis for identifying candidate outer membrane proteins from Borrelia spirochetes

Abstract

BackgroundSimilar to Gram-negative organisms, Borrelia spirochetes are dual-membrane organisms with both an inner and outer membrane. Although the outer membrane contains integral membrane proteins, few of the borrelial outer membrane proteins (OMPs) have been identified and characterized to date. Therefore, we utilized a consensus computational network analysis to identify novel borrelial OMPs.ResultsUsing a series of computer-based algorithms, we selected all protein-encoding sequences predicted to be OM-localized and/or to form β-barrels in the borrelial OM. Using this system, we identified 41 potential OMPs from B. burgdorferi and characterized three (BB0838, BB0405, and BB0406) to confirm that our computer-based methodology did, in fact, identify borrelial OMPs. Triton X-114 phase partitioning revealed that BB0838 is found in the detergent phase, which would be expected of a membrane protein. Proteolysis assays indicate that BB0838 is partially sensitive to both proteinase K and trypsin, further indicating that BB0838 is surface-exposed. Consistent with a prior study, we also confirmed that BB0405 is surface-exposed and associates with the borrelial OM. Furthermore, we have shown that BB0406, the product of a co-transcribed downstream gene, also encodes a novel, previously uncharacterized borrelial OMP. Interestingly, while BB0406 has several physicochemical properties consistent with it being an OMP, it was found to be resistant to surface proteolysis. Consistent with BB0405 and BB0406 being OMPs, both were found to be capable of incorporating into liposomes and exhibit pore-forming activity, suggesting that both proteins are porins. Lastly, we expanded our computational analysis to identify OMPs from other borrelial organisms, including both Lyme disease and relapsing fever spirochetes.ConclusionsUsing a consensus computer algorithm, we generated a list of candidate OMPs for both Lyme disease and relapsing fever spirochetes and determined that three of the predicted B. burgdorferi proteins identified were indeed novel borrelial OMPs. The combined studies have identified putative spirochetal OMPs that can now be examined for their roles in virulence, physiology, and disease pathogenesis. Importantly, the studies described in this report provide a framework by which OMPs from any human pathogen with a diderm ultrastructure could be cataloged to identify novel virulence factors and vaccine candidates.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0762-z) contains supplementary material, which is available to authorized users.

Outer Membrane Proteins BB0405 and BB0406 Are Immunogenic, but Only BB0405 Is Required for Borrelia burgdorferi Infection

“…The genes encoding these OMPs were recently shown to be cotranscribed (32,35), suggesting that these OMPs would be coexpressed. However, in this study, we observed a difference in the temporal antibody response to the individual BB0405 and BB0406 OMPs, suggesting that they may not be coordinately expressed during infection.…”

“…Outer membrane protein BB0405 was initially identified in a screen for outer surface proteins (34) and was more recently characterized for its physicochemical properties (32) and role in B. burgdorferi virulence (35). Additional studies on BB0405 orthologs of other borrelial genospecies have indicated that this OMP binds human factor H (36,37), suggesting that BB0405 and its orthologs play a role in serum resistance.…”

“…Despite these limitations, seven membrane-spanning OMPs unique to B. burgdorferi have been identified. Five of these OMPs, P66, P13, DipA, Lmp1, and BB0172, have been shown to form pores and/or act as adhesins (21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31), while the remaining two, BB0405 and BB0406, are paralogs that have been shown to form pores in large unilamellar vesicles (32,33).…”

“…Additional studies on BB0405 orthologs of other borrelial genospecies have indicated that this OMP binds human factor H (36,37), suggesting that BB0405 and its orthologs play a role in serum resistance. While the physicochemical properties of BB0406 were recently examined, revealing that this OMP can form a pore (32), the role of BB0406 in borrelial virulence has not yet been explored. Therefore, to determine if BB0406 is important for infection and serum resistance, similarly to its paralog BB0405, we further examined both OMPs for their role(s) in animal infection, factor H binding activity, and serum resistance.…”

Delineating Surface Epitopes of Lyme Disease Pathogen Targeted by Highly Protective Antibodies of New Zealand White Rabbits

“…In addition to Lmp1, P66, and enolase, two other chromosomally encoded proteins (outer membrane proteins), BB0838 and BB0465, were also mapped by PA peptides. BB0838, mapped by p202B71, is a 120-kDa outer membrane protein which was experimentally demonstrated to be surface exposed (99). The fact that no transposon was inserted in the bb0838 gene upon global transposon mutagenesis (100, 101) strongly suggests that BB0838 plays an essential role in the life cycle of B. burgdorferi.…”