Initiating the Classical Pathway: IgG recruitment, Hexamerization, and C1q binding in Real-Time

Complement activation is triggered by the formation of Immunoglobulin (IgG) hexamers on the surface of pathogens, tumors, or autoantigenic cells (1–3). How IgG binding to surface antigens induces the dynamic assembly of IgG hexamers that recruit and activate C1, the first component of complement, is poorly understood. Here, we employed high-speed atomic force microscopy (4–7) to directly visualize dynamic IgG binding and hexamer formation on antigenic lipid bilayer membranes, the subsequent binding of C1q, and enhanced surface IgG hexamerization by mutations promoting interactions between neighboring antibodies. With single-molecule force spectroscopy and quartz crystal microbalance we further characterized the molecular interactions by determining chemical rate constants and energies. Our data suggest that antigen recognition by IgGs may nucleate subsequent oligomerization through IgG recruitment from solution and, depending on the valency of its binding state, through lateral collisions. This finally leads to stable IgG hexamers competent of tightly binding C1q and initiating the classical complement cascade.