HDX-MS and MD Simulations Provide Evidence for Stabilization of the IgG1-FcγRIa (CD64a) Immune Complex Through Intermolecular Glycoprotein Bonds

J Mol Biol. 2022 Jan 30;434(2):167391. doi: 10.1016/j.jmb.2021.167391. Epub 2021 Dec 8.

Abstract

Previous reports present different models for the stabilization of the Fc-FcγRI immune complex. Although accord exists on the importance of L235 in IgG1 and some hydrophobic contacts for complex stabilization, discord exists regarding the existence of stabilizing glycoprotein contacts between glycans of IgG1 and a conserved FG-loop (171MGKHRY176) of FcγRIa. Complexes formed from the FcγRIa receptor and IgG1s containing biantennary glycans with N-acetylglucosamine, galactose, and α2,6-N-acetylneuraminic terminations were measured by hydrogen-deuterium exchange mass spectrometry (HDX-MS), classified for dissimilarity with Welch's ANOVA and Games-Howell post hoc procedures, and modeled with molecular dynamics (MD) simulations. For each glycoform of the IgG1-FcγRIa complex peptic peptides of Fab, Fc and FcγRIa report distinct H/D exchange rates. MD simulations corroborate the differences in the peptide deuterium content through calculation of the percent of time that transient glycan-peptide bonds exist. These results indicate that stability of IgG1-FcγRIa complexes correlate with the presence of intermolecular glycoprotein interactions between the IgG1 glycans and the 173KHR175 motif within the FG-loop of FcγRIa. The results also indicate that intramolecular glycan-protein bonds stabilize the Fc region in isolated and complexed IgG1. Moreover, HDX-MS data evince that the Fab domain has glycan-protein binding contacts within the IgG1-FcγRI complex.

Keywords: glycosylation; immune receptor; membrane protein structure; monoclonal antibody; similarity.

MeSH terms

  • Antibodies, Monoclonal / chemistry
  • Antigen-Antibody Complex / chemistry*
  • Antigen-Antibody Complex / metabolism
  • Galactose
  • Glycoproteins / chemistry*
  • Glycoproteins / metabolism
  • Hydrogen Deuterium Exchange-Mass Spectrometry / methods*
  • Immunoglobulin G / chemistry*
  • Membrane Proteins / chemistry
  • Molecular Dynamics Simulation*
  • Peptides / chemistry
  • Peptides / metabolism
  • Polysaccharides
  • Protein Binding
  • Receptors, IgG / chemistry*

Substances

  • Antibodies, Monoclonal
  • Antigen-Antibody Complex
  • FCGR1A protein, human
  • Glycoproteins
  • Immunoglobulin G
  • Membrane Proteins
  • Peptides
  • Polysaccharides
  • Receptors, IgG
  • Galactose