Abstract
The finding that the antibody (Ab) constant (C) region can influence fine specificity suggests that isotype switching contributes to the generation of Ab diversity and idiotype restriction. Despite the centrality of this observation for diverse immunological effects such as vaccine responses, isotype-restricted antibody responses, and the origin of primary and secondary responses, the molecular mechanism(s) responsible for this phenomenon are not understood. In this study, we have taken a novel approach to the problem by probing the paratope with (15)N label peptide mimetics followed by NMR spectroscopy and fluorescence emission spectroscopy. Specifically, we have explored the hypothesis that the C region imposes conformational constraints on the variable (V) region to affect paratope structure in a V region identical IgG(1), IgG(2a), IgG(2b), and IgG(3) mAbs. The results reveal isotype-related differences in fluorescence emission spectroscopy and temperature-related differences in binding and cleavage of a peptide mimetic. We conclude that the C region can modify the V region structure to alter the Ab paratope, thus providing an explanation for how isotype can affect Ab specificity.
Highlights
The mechanism by which antibody constant region alters fine specificity is unknown
The finding that the antibody (Ab) constant (C) region can influence fine specificity suggests that isotype switching contributes to the generation of Ab diversity and idiotype restriction
We have taken a novel approach to the problem by probing the paratope with 15N label peptide mimetics followed by NMR spectroscopy and fluorescence emission spectroscopy
Summary
The mechanism by which antibody constant region alters fine specificity is unknown. Results: Different constant regions were found to change electronic and chemical properties of the antigen-binding site. Previous studies done in our lab using four murine mAb isotypes, IgG1, IgG2a, IgG2b, and IgG3 suggest that the C region imposes structural constraints on the V region that alter its structure and/or ability to undergo a conformational change upon Ag binding [7, 15,16,17,18] They are identical in sequence, the V regions of this group of mAbs may have secondary structures capable of different interactions that manifest themselves as changed fine specificity. This view is supported by several lines of evidence. Our results provide direct experimental support for the notion that the C domain can affect antibody fine specificity by influencing the chemical and electronic environment of the Ab paratope
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