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Quantitative analysis of the interaction strength and dynamics of human IgG4 half molecules by native mass

Rebecca J Rose1, Aran F Labrijn, Ewald T J van den Bremer

  • 1Biomolecular Mass Spectrometry and Proteomics Group, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.

Structure (London, England : 1993)
|September 7, 2011
PubMed

Insights

Native mass spectrometry quantified antibody half molecule homodimerization in human IgG4. This revealed critical CH3-CH3 interface interactions essential for dimeric structure and biological function.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Immunology

Background:

  • Antibody half molecules (HL) form dimers through noncovalent interactions.
  • Understanding these interactions is crucial for antibody structure and function.
  • Human IgG4 (IgG4Δhinge) provides a model for studying HL homodimerization.

Purpose of the Study:

  • To investigate the noncovalent interactions in IgG4 half molecule homodimerization.
  • To quantify the dissociation constant (K(D)) of this interaction.
  • To elucidate the role of the CH3-CH3 interface in stabilizing the dimeric structure.

Main Methods:

  • Native mass spectrometry (MS) to analyze protein complexes.
  • Concentration-dependent analysis of monomer and dimer species.
  • Site-directed mutagenesis to probe specific residues.
  • Time-resolved MS for kinetic analysis.

Main Results:

  • Apparent dissociation constants (K(D)) for HL homodimerization were determined.
  • Key residues at the CH3-CH3 interface were identified and quantified.
  • K(D) values ranged from 10(-10) to 10(-4) M.
  • Noncovalent interactions were shown to be critical for full-length IgG4 dimeric structure.
  • Kinetics of IgG4 HL exchange were measured.

Conclusions:

  • Native MS provides quantitative insights into antibody half molecule homodimerization.
  • The CH3-CH3 interface plays a vital role in IgG4 structural integrity.
  • This study links local structural features to the biological properties of IgG4.

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