What question did this study set out to answer?

To elucidate the mechanistic basis of FcγRIIa selectivity by monoclonal antibody IV.3 through molecular simulations.

February 21, 2026

BPS2026 – Mechanistic basis of FcγRIIa selective recognition by monoclonal antibody IV.3 revealed by molecular simulations

Key Points

To elucidate the mechanistic basis of FcγRIIa selectivity by monoclonal antibody IV.3 through molecular simulations.
Combined atomistic molecular dynamics and alchemical free-energy simulations of the FcγRIIa-IV.3 complex.
Detailed thermodynamic cycle analysis considering mutations H134R, L135S, and H134R/L135S.
Single mutations modestly altered affinity, while the double mutant synergistically destabilized binding.
Simulations indicated Y122 flips to encapsulate R134, forming salt bridges and cation-π interactions, pivotal for specificity.

Abstract

Engagement of the platelet FcγIIa receptor ( FcγRIIa) by immune complexes leads to integrin αIIbβ3 activation and platelet aggregation, driving thrombo-inflammatory responses in infection, autoimmunity, and cardiovascular disease. As this engagement does not contribute to hemostasis, FcγRIIa selective inhibition could suppress pathological platelet activation without impairing normal clotting. The murine monoclonal antibody (mAb) IV.3 is widely used to probe FcγRIIa function due to its high specificity over the closely related FcγRIIb. Its epitope includes residues 132–137, including the FcγRIIa-specific H/R134 polymorphism (rs1801274) and L135 (R134/S135 in FcγRIIb), but the mechanistic basis of selectivity remains unclear. We combined atomistic molecular dynamics and alchemical free-energy simulations of the cryo-electron microscopy structure of the FcγRIIa-IV.3 complex to elucidate the binding mechanism. A thermodynamic cycle encompassing H134R, L135S, and the H134R/L135S double mutant (mimicking FcγRIIb) revealed that single mutations modestly altered affinity, whereas H134R/L135S synergistically destabilized binding. These results align with surface plasmon resonance measurements of FcγRIIa and FcγRIIb binding to immobilized IV.3. Protein-protein interaction profiling of our simulations indicated that L135 contributes to critical hydrophobic contacts during encounter complex formation, while the H/R134 substitution introduces favorable electrostatic and cation-π interactions, rationalizing modest affinity gains. Strikingly, FcγRIIa-R134 simulations revealed a conformational rearrangement in which Y122 flips to encapsulate R134, forming salt bridges with D119 and D123 and cation-π interactions with Y122, Y73, and Y51, consistent with AlphaFold2-predicted states. This gating-like rearrangement between Y122 and Y51 emerges as a previously unrecognized feature of FcγRIIa-IV.3 recognition. Together, these findings support a molecular mechanism in which hydrophobic stabilization by L135 primes a loop rearrangement, enabling H/R134-dependent electrostatic and aromatic interactions to fine-tune specificity. These insights rationalize IV.3 selectivity for FcγRIIa and provide a framework for designing next-generation FcγRIIa-targeted therapeutics.

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