Biophysical characterization of human complement component 1q (C1q) binding to IgG oligomers

The human immune system consists of powerful defense mechanisms that recognize and eliminate pathological agents from our bodies. One branch of these defenses is the classical complement system, which utilizes the highly specific binding of antibodies to their antigens to initiate a rapid cascade that ultimately leads to cell lysis. Cascade activation is mediated by C1q, a soluble hexavalent complex that recognizes antibodies on the cell surface, but the exact mechanism of C1q binding and activity remains unclear1. A number of studies have shown that antibody hexamers provide ideal binding sites for complement2,3. Our previous work has revealed the IgG hexamerization process to be a complex series of events that requires antigenic surfaces. We have shown that complement-enhancing mutations at the Fc region directly amplify hexamerization, and that efficient complement activation requires C1q to recognize at least four binding sites on IgG oligomers4. Here we investigate the C1q binding process itself using High-Speed Atomic Force Microscopy (HS-AFM) and Quartz Crystal Microbalance (QCM). We report the dynamics of C1q binding to a range of IgG mutants at various surface densities and states of oligomerization, and provide rates of association as well as C1q recruitment and retention efficacies not resolved by traditional complement assays. 1. Gaboriaud, C., Ling, W. L., Thielens, N. M., Bally, I. & Rossi, V. Deciphering the Fine Details of C1 Assembly and Activation Mechanisms: “Mission Impossible”? Front. Immunol. 5, (2014). 2. Diebolder, C. A. et al. Complement Is Activated by IgG Hexamers Assembled at the Cell Surface. Science 343, 1260–1263 (2014). 3. Jong, R. N. de et al. A Novel Platform for the Potentiation of Therapeutic Antibodies Based on Antigen-Dependent Formation of IgG Hexamers at the Cell Surface. PLOS Biology 14, e1002344 (2016). 4. Strasser, J. et al. Elucidation of the IgG oligomerization process that triggers classical pathway complement activation. Submitted