The increased hinge flexibility of an IgG1-IgG3 hybrid monoclonal enhances Fc-mediated protection against group A streptococci

Antibodies are central to the immune response against microbes. We have previously generated a protective IgG1 monoclonal antibody targeting the M protein, a critical virulence factor of Streptococcus pyogenes. Here, we generated this antibody in all human IgG subclasses and evaluated their function. Despite significantly reduced binding, the IgG3 subclass antibody demonstrated remarkably enhanced opsonic function. We hypothesized that increased Fc flexibility could explain this improved efficacy. We engineered a hybrid IgG subclass antibody, IgGh, containing the backbone of IgG1 with the hinge of IgG3, leaving the Fabs unchanged. The IgGh maintained a similar binding ability as IgG1 while gaining the strong opsonic function seen with IgG3. Molecular dynamics simulations of the different antibodies showed altered IgG Fab-antigen interactions, reflecting the differences observed in affinity. More importantly, when the antibodies were bound to the antigen, the simulations showed that the Fc of both IgGh and IgG3 exhibited extensive movement in 3D space relative to the M protein. The increased flexibility of IgGh directly translated to enhanced opsonic function and significantly increased the protection against infection with Streptococcus pyogenes in mice. Our findings demonstrate how altering Fc flexibility can improve Fc-mediated opsonic function and how modifications in the constant domain can regulate Fab-antigen interactions. In addition, the enhanced in vivo function of a more flexible IgG provides new therapeutic opportunities for monoclonal antibodies. One sentence summary Antibody Fc flexibility in 3D space correlates with efficient Fc-mediated phagocytosis of streptococci ### Competing Interest Statement HK, WB, LH, OS, JM, LM, PN have a patent regarding Ab25 and AI, PN has filed a patent related to the findings in this manuscript.