PhD (Doctor of Philosophy)
Date of Award
Committee Chair or Co-Chairs
Patrick Bradshaw, Diego Rodriguez Gil, Jennifer Hall, Cecilia McIntosh, Christopher Pritchett
C-reactive protein (CRP) is a part of the innate immune system, is synthesized in the liver, its blood level increases in inflammatory states, and it binds to Streptococcus pneumoniae. The conformation of CRP is altered under conditions mimicking an inflammatory milieu and this non-native CRP also binds to immobilized/aggregated/pathogenic proteins. Experiments in mice have revealed that one of the functions of CRP is to protect against pneumococcal infection. For protection, CRP must be injected into mice within two hours of administering pneumococci, thus, CRP is protective against early-stage infection but not against late-stage infection. It is unknown how CRP protects or why CRP does not protect against late-stage infection. The hypotheses are that the protection requires complement activation by CRP-pneumococci complexes and that CRP cannot protect if pneumococci have time to recruit complement inhibitor factor H on their surface to become complement attack-resistant. To test these hypotheses, we generated CRP mutants by site-directed mutagenesis: a mutant that binds to pneumococci but does not activate complement and a mutant that binds to immobilized factor H. We found that mutant CRP incapable of activating complement was not protective against infection and that mutant CRP capable of binding to factor H was protective against both early and late stage infections. Additional experiments showed that CRP enhances the effects of the antibiotic clarithromycin in reducing bacteremia in infected mice. Moreover, we observed that mutant CRP capable of binding to factor H bound to several proteins immobilized on plastic, suggesting that CRP recognizes a pattern, probably an amyloid-like structure, on immobilized proteins. Indeed, mutant CRP, after binding to amyloid b peptides, prevented the formation of pathogenic amyloid fibrils. Lastly, employing a hepatic cell line, we investigated the mechanism of CRP expression in response to pro-inflammatory cytokines. We found that the transcription factor C/EBPb and two C/EBP-binding sites on the CRP promoter were critical for inducing CRP expression. We conclude that complement activation is necessary for CRP-mediated protection against infection, that CRP functions in two structural conformations, that CRP and clarithromycin act synergistically, that CRP has anti-amyloidogenic properties, and the increased CRP expression requires C/EBPb.
Dissertation - embargo
Ngwa, Donald, "A Novel Mode of Action of C-reactive Protein in Protecting Against Streptococcus pneumoniae Infection and Synergy with Antibiotics" (2020). Electronic Theses and Dissertations. Paper 3752. https://dc.etsu.edu/etd/3752
Copyright by the authors.
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