Abstract
Antibodies are secreted proteins that are crucial to recognition of pathogens by the immune system and are also efficient pharmaceuticals. The affinity and specificity of target recognition can increase remarkably through avidity effects, when the antibody can bind a multivalent antigen through more than one epitope simultaneously. A key goal of antibody engineering is thus to optimize avidity, but little is known about the nanoscale spatial dependence of avidity in antibodies. Here, we develop a set of anti-parallel coiled-coils spanning from 7 to 20 nm and validate their structure using biophysical techniques. We use the coiled-coils to control the spacing between two epitopes, and measure how antigen spacing affects the stability of the bivalent antibody:antigen complex. We find a maximal avidity enhancement at a spacing of 13 nm. In contrast to recent studies, we find the avidity to be relatively insensitive to epitope spacing near the avidity maximum as long as it is within the spatial tolerance of the antibody. We thus only see a ~ twofold variation of avidity in the range from 7 to 20 nm. The coiled-coil systems developed here may prove a useful protein nanocaliper for profiling the spatial tolerance and avidity profile of bispecific antibodies.
Highlights
Antibodies are secreted proteins that are crucial to recognition of pathogens by the immune system and are efficient pharmaceuticals
Antibodies are proteins secreted by the immune system that detect and neutralize foreign molecules
We know little about how avidity in antibodies depends on the spatial arrangement of epitopes, which prevents the rational use of avidity in antibody engineering
Summary
Antibodies are secreted proteins that are crucial to recognition of pathogens by the immune system and are efficient pharmaceuticals. Rare crystal structures of full-length antibodies suggested that a fully extended hinge (Fig. 1A) would allow a distance up to 9 nm between the binding site and the first disulfide bond in the hinge. Bivalent binding occurs in two steps: An initially monovalent association is followed by an intra-complex ring-closing reaction to form the cyclical complex (Fig. 1B).
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