Isothermal titration calorimetry reveals differential binding thermodynamics of variable region-identical antibodies differing in constant region for a univalent ligand.
The classical view of immunoglobulin molecules posits two functional domains defined by the variable (V) and constant (C) regions, which are responsible for antigen binding and antibody effector functions, respectively. These two domains are thought to function independently. However, several lines of evidence strongly suggest that C region domains can affect the specificity and affinity of an antibody for its antigen (Ag), independent of avidity-type effects. In this study, we used isothermal titration calorimetry to investigate the thermodynamic properties of the interactions of four V region-identical monoclonal antibodies with a univalent peptide antigen. Comparison of the binding of IgG1, IgG2a, IgG2b, and IgG3 with a 12-mer peptide mimetic of Cryptococcus neoformans polysaccharide revealed a stoichiometry of 1.9-2.0 with significant differences in thermodynamic binding parameters. Binding of this peptide to the antibodies was dominated by favorable entropy. The interaction of these antibodies with biotinylated peptides manifested greater enthalpy than for native peptides indicating that biotin labeling affected the types of Ag-Ab complexes formed. Our results provide unambiguous thermodynamic evidence for the notion that the C region can affect the interaction of the V region with an Ag.
Dam
Tarun K
TK
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461, USA. tdam@aecom.yu.edu
Torres
Marcela
M
Brewer
C Fred
CF
Casadevall
Arturo
A
D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry* / Q000276:immunology*; D019921:Biotinylation; D002151:Calorimetry; D018360:Crystallography, X-Ray; D008024:Ligands; D010455:Peptides / Q000276:immunology / Q000378:metabolism; D011485:Protein Binding; D013696:Temperature; D013816:Thermodynamics; D017186:Titrimetry
D000906:Antibodies; D008024:Ligands; D010455:Peptides