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Two high-affinity monoclonal IgG2a antibodies with differing thermodynamic stability demonstrate distinct
Z I Kravchuk1, A A Chumanevich, A P Vlasov
1Department of Protein Chemistry, Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, Minsk.
Journal of Immunological Methods
|October 17, 1998
Summary
Two monoclonal antibodies (mAbs) targeting human spleen ferritin show different protein A-binding changes upon antigen interaction. This difference is linked to structural variations and thermodynamic stability, impacting antibody function.
Area of Science:
- Immunology
- Biochemistry
- Structural Biology
Background:
- Monoclonal antibodies (mAbs) are crucial tools in research and therapeutics.
- Protein A is widely used for antibody purification due to its affinity for the Fc region of immunoglobulins.
- Understanding how antigen binding affects antibody-protein A interactions is important for antibody engineering.
Purpose of the Study:
- To investigate the differential effects of antigen binding on the protein A-binding affinity of two IgG2a monoclonal antibodies (G10 and F11) targeting human spleen ferritin.
- To explore the structural and thermodynamic basis for these observed differences.
Main Methods:
- Characterization of human spleen ferritin binding affinity for G10 and F11.
- Assessment of protein A-binding affinity before and after antigen binding.
- Differential scanning calorimetry (DSC) to determine thermodynamic stability (enthalpy and Gibb's free energy of unfolding).
Main Results:
- Both G10 and F11 bind human spleen ferritin with similar affinities.
- Antigen binding modulates protein A affinity differently: G10's affinity increases, while F11's affinity decreases significantly.
- Differential scanning calorimetry revealed lower structural energetics for F11 compared to G10, suggesting a lack of a specific folding unit influencing binding site interactions.
Conclusions:
- Subclass-independent structural variations can modulate interactions between antibody recognition sites.
- Thermodynamic stability plays a role in the functional consequences of antigen-antibody complex formation.
- This study demonstrates how conformational changes upon antigen binding can lead to distinct protein A-binding behaviors in antibodies of the same subclass.