Contribution of IgG Subclass and Heavy Chain Constant Region Domains to Affinity and Protective Activity of mAbs Specific to the £^DPGA Capsule of Bacillus anthracis.
AdvisorKozel, Thomas R
Biochemistry and Molecular Biology
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Bacillus anthracis, the causative agent of anthrax, is surrounded by an antiphagocytic capsule composed of poly-£^-D-glutamic acid (£^DPGA). Antibodies directed against the £^DPGA capsule are opsonic and protective, which makes the capsule an attractive target for vaccine development. However, antibodies of different IgG subclasses differ in their ability to activate effector functions, including activation of complement and phagocytosis. In addition, recent studies found differences in affinity and protective activity between variable region identical IgG subclass switch monoclonal antibodies (mAbs) directed against microbial capsules or outer membrane proteins. We recently generated six mAbs against the £^DPGA capsule and found that only mAbs of IgG3 subclass were protective in a murine model of inhalational anthrax. In order to investigate the role of IgG subclass to protective activity against anthrax, the goal of the study was to generate a full IgG subclass switch family (IgG3 ,,³ IgG1 ,,³ IgG2b ,,³ IgG2a) from two of our £^DPGA mAbs, F26G3 and F24F2, and evaluate these mAbs for their biological and protective activities. Our results showed that only mAbs of IgG3 subclass were protective in vivo, whereas the IgG1, IgG2b, or IgG2a subclass switch mAbs failed to protect. Protective activity of mAbs correlated with high affinity and ability to activate complement and opsonophagocytosis in vitro. These results suggested that the IgG heavy chain constant region (CH) domains contribute to antibody-antigen interaction and ability to protect. To further examine the role of IgG CH domains to antibody function, we generated CH swap mAbs from our protective F26G3 IgG3 and non-protective IgG2b mAbs. Our results suggested a major contribution for the CH2 and CH3 domains of IgG3 for antibody affinity, antibody-antigen interaction and protective activity. Taken together, our results showed that IgG heavy chain contributes to antibody affinity and antibody-antigen interaction, which further affects the protective activity of the anti-£^DPGA mAbs.