Unraveling the Macromolecular Pathways of IgG Oligomerization and Complement Activation on Antigenic Surfaces

Abstract

IgG antibodies play a central role in protection against pathogens by their ability to alert and activate the innate immune system. Here, we show that IgGs assemble into oligomers on antigenic surfaces through an ordered, Fc domain-mediated process that can be modulated by protein engineering. Using high-speed atomic force microscopy, we unraveled the molecular events of IgG oligomer formation on surfaces. IgG molecules were recruited from solution although assembly of monovalently binding molecules also occurred through lateral diffusion. Monomers were observed to assemble into hexamers with all intermediates detected, but in which only hexamers bound C1. Functional characterization of oligomers on cells also demonstrated that C1 binding to IgG hexamers was a prerequisite for maximal activation, whereas tetramers, trimers, and dimers were mostly inactive. We present a dynamic IgG oligomerization model, which provides a framework for exploiting the macromolecular assembly of IgGs on surfaces for tool, immunotherapy, and vaccine design.

On-Membrane Dynamic Interplay between Anti-GM1 IgG Antibodies and Complement Component C1q

“…In recent studies, the binding of C1q with IgG oligomers was studied, most notably by employing artificially hexamerized IgG antibodies [11][12][13][14]. It has been reported that IgG molecules have the potential ability to form hexamers in crystal or on mica surface under certain conditions [14,24].…”

“…Our STD data obtained by using a GM2 tetrasaccharide also confirmed that the interaction was significantly compromised by the removal of the outer galactose residue. In the subsequent experiments, we characterized the interaction of GB2 with GM1 in membrane environments (Figure 1c 13 C HSQC spectrum (black) and the 1 H-13 C STD-HSQC spectrum (red) of the 2-trimethylsilylethyl derivatives of the GM2 tetrasaccharide (10 equiv.) with 50 μM GB2.…”

“…These results indicate that antigen binding promotes the hexameric ring formation of the GB2 antibodies on the membrane. 13 C HSQC spectrum (black) and the 1 H-13 C STD-HSQC spectrum (red) of the 2-trimethylsilylethyl derivatives of the GM2 tetrasaccharide (10 equiv.) with 50 µM GB2.…”

“…Next, we attempted to address the pathological relevance of the antigen-dependent IgG hexamer formation. We examined the potential interaction of the IgG hexameric ring with C1q using HS-AFM, because accumulating data have indicated that C1q preferentially binds to IgG oligomers [11][12][13][14]. HS-AFM could clearly visualize dynamic structures of free C1q molecules on mica surface treated with 3-aminopropyltriethoxysilane, exhibiting six highly mobile globular heads tethered to the central stem, whereas C1q molecules were not clearly observable on the membranes because of rapid diffusion due to weak interaction with the membrane surface (Figure 3a…”

Antibody barriers to going viral

“…Thus, for example, two Abs, nominally to the same epitope, may well have somewhat different angles of approach to their target and may present a somewhat different array of Fcs to an effector cell with potential differences in effector function. For example, the importance of Ab arrays is suggested to be critical in complement triggering (Diebolder et al, 2014;Strasser et al, 2019). Furthermore, this discussion refers to mAbs.…”