Abstract
Factor H binding protein (FHbp) is a virulence factor used by meningococci to evade the host complement system. FHbp elicits bactericidal antibodies in humans and is part of two recently licensed vaccines. Using human complement Factor H (FH) transgenic mice, we previously showed that binding of FH decreased the protective antibody responses to FHbp vaccination. Therefore, in the present study we devised a library-based method to identify mutant FHbp antigens with very low binding of FH. Using an FHbp sequence variant in one of the two licensed vaccines, we displayed an error-prone PCR mutant FHbp library on the surface of Escherichia coli. We used fluorescence-activated cell sorting to isolate FHbp mutants with very low binding of human FH and preserved binding of control anti-FHbp monoclonal antibodies. We sequenced the gene encoding FHbp from selected clones and introduced the mutations into a soluble FHbp construct. Using this approach, we identified several new mutant FHbp vaccine antigens that had very low binding of FH as measured by ELISA and surface plasmon resonance. The new mutant FHbp antigens elicited protective antibody responses in human FH transgenic mice that were up to 20-fold higher than those elicited by the wild-type FHbp antigen. This approach offers the potential to discover mutant antigens that might not be predictable even with protein structural information and potentially can be applied to other microbial vaccine antigens that bind host proteins.
Highlights
Binding of complement regulators is a common strategy that microbes use to subvert the host immune system
Meningococcal Factor H binding protein (FHbp) expressed with its native signal sequence was detected on the surface of E. coli by flow cytometry using a mixture of two murine anti-FHbp monoclonal antibodies, JAR 41 [29] and mAb502 [30] (Fig 1A, solid blue line)
The R41S mutant did not bind Factor H (FH), it was present on the bacterial surface since it was detected by the control murine mAb (Fig 1A, dashed green line)
Summary
Binding of complement regulators is a common strategy that microbes use to subvert the host immune system. It has been proposed, that vaccines targeting the microbial ligands for host complement regulators would elicit antibodies that defeat this evasion mechanism [1]. The effectiveness of microbial proteins that bind host complement regulators as vaccine antigens might be hampered by binding of the host protein. Modification of the microbial antigen so as not to bind the complement regulator might increase immunogenicity [1]. Several microbial vaccine antigens that bind complement regulators, including HIV gp120.
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