Dynamic Views of the Fc Region of Immunoglobulin G Provided by Experimental and Computational Observations.

Saeko Yanaka,Tadashi Satoh,Hisashi Okumura,Satoru G Itoh,Hirokazu Yagi,Yohei Miyanoiri,Koichi Kato,Masaaki Sugiyama,Rintaro Inoue,Takumi Yamaguchi,Rina Yogo

doi:10.3390/antib8030039

Abstract

The Fc portion of immunoglobulin G (IgG) is a horseshoe-shaped homodimer, which interacts with various effector proteins, including Fcγ receptors (FcγRs). These interactions are critically dependent on the pair of N-glycans packed between the two CH2 domains. Fucosylation of these N-glycans negatively affects human IgG1-FcγRIIIa interaction. The IgG1-Fc crystal structures mostly exhibit asymmetric quaternary conformations with divergent orientations of CH2 with respect to CH3. We aimed to provide dynamic views of IgG1-Fc by performing long-timescale molecular dynamics (MD) simulations, which were experimentally validated by small-angle X-ray scattering and nuclear magnetic resonance spectroscopy. Our simulation results indicated that the dynamic conformational ensembles of Fc encompass most of the previously reported crystal structures determined in both free and complex forms, although the major Fc conformers in solution exhibited almost symmetric, stouter quaternary structures, unlike the crystal structures. Furthermore, the MD simulations suggested that the N-glycans restrict the motional freedom of CH2 and endow quaternary-structure plasticity through multiple intramolecular interaction networks. Moreover, the fucosylation of these N-glycans restricts the conformational freedom of the proximal tyrosine residue of functional importance, thereby precluding its interaction with FcγRIIIa. The dynamic views of Fc will provide opportunities to control the IgG interactions for developing therapeutic antibodies.

Highlights

Antibodies play pivotal roles in the immune system as multifunctional glycoproteins, coupling between antigen recognition and effector functions, as typified by immunoglobulin G (IgG)
The root mean square fluctuation (RMSF) for each amino acid Cα atom of IgG1-Fc was calculated from 3,200 conformers extracted from each of the eight production runs, which were superimposed by the Cα atoms, yielding an average structure
The great majority of the IgG1-Fc crystal structures deposited in Protein Data Bank (PDB) exhibit asymmetric quaternary structures even in uncomplexed states, with few exceptions, for example, 5IW3 with a crystallographic two-fold axis

Summary

Introduction

Antibodies play pivotal roles in the immune system as multifunctional glycoproteins, coupling between antigen recognition and effector functions, as typified by immunoglobulin G (IgG). The Fc portion of IgG interacts with various proteins other than the effector proteins, such as staphylococcal protein A and streptococcal protein G [5]. This versatile functionality of IgG and the other classes of antibodies is believed to be attributed to their structural flexibility and plasticity [6,7]. Antibodies have modular structures, in which the Ig-fold domains as building blocks are connected through flexible linkers. The CH 1 and CH 2 domains are separated by the hinge region, which possesses significant degrees of freedom for internal flexibility

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Conclusion