Recognition of Human IgG1 by Fcγ Receptors: Structural Insights from Hydrogen-Deuterium Exchange and Fast Photochemical Oxidation of Proteins Coupled with Mass Spectrometry

Biochemistry. 2019 Feb 26;58(8):1074-1080. doi: 10.1021/acs.biochem.8b01048. Epub 2019 Feb 6.

Abstract

Antibody-dependent cell-mediated cytotoxicity (ADCC) is an effector function of immunoglobulins (IgGs) involved in the killing of target cells by a cytotoxic effector cell. Recognition of IgG by Fc receptors expressed on natural killer cells, mostly FcγIII receptors (FcγRIII), underpins the ADCC mechanism, thus motivating investigations of these interactions. In this paper, we describe the combination of hydrogen-deuterium exchange and fast photochemical oxidation of proteins (FPOP) coupled with mass spectrometry to study the interactions of the human IgG1/FcγRIII complex. Using these orthogonal approaches, we identified critical peptide regions and residues involved in the recognition of IgG1 by FcγRIII. The footprinting results are consistent with the previously published crystal structure of the IgG1 Fc/FcγRIII complex. Additionally, our FPOP results reveal the conformational changes in the Fab domain upon binding of the Fc domain to FcγRIII. These data demonstrate the value of footprinting as part of a comprehensive toolbox for identifying the changes in the higher-order structure of therapeutic antibodies in solution.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibody-Dependent Cell Cytotoxicity
  • Deuterium / chemistry*
  • Deuterium Exchange Measurement*
  • GPI-Linked Proteins / chemistry
  • GPI-Linked Proteins / metabolism
  • Humans
  • Hydrogen / chemistry*
  • Immunoglobulin Fc Fragments / chemistry
  • Immunoglobulin Fc Fragments / metabolism
  • Immunoglobulin G / chemistry*
  • Immunoglobulin G / metabolism
  • Mass Spectrometry / methods*
  • Models, Molecular
  • Oxidation-Reduction
  • Receptors, IgG / chemistry*
  • Receptors, IgG / metabolism

Substances

  • FCGR3A protein, human
  • FCGR3B protein, human
  • GPI-Linked Proteins
  • Immunoglobulin Fc Fragments
  • Immunoglobulin G
  • Receptors, IgG
  • Hydrogen
  • Deuterium