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. The molecular pathway of how IgG binding to surface antigens initiates and facilitates hexamerization remains poorly understood. Here, we employed high-speed atomic force microscopy to directly visualize dynamic IgG binding and hexamer formation on antigenic lipid bilayer membranes and subsequent binding of C1q– the first component of complement- in real-time and under physiological conditions. With single-molecule force spectroscopy and quartz crystal microbalance we further characterized the molecular interactions by determining chemical rate constants and energies. Our data outline a process in which antigen recognition by IgGs nucleates subsequent oligomerization through IgG recruitment from solution or via lateral collisions. This finally leads to stable IgG hexamers competent of tightly binding C1q and thus initiates the classical complement pathway.