Affinity maturation increases the stability and plasticity of the Fv domain of anti-protein antibodies |
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Authors: | Acierno Juan Pablo Braden Bradford C Klinke Sebastián Goldbaum Fernando A Cauerhff Ana |
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Affiliation: | 1 Fundación Instituto Leloir, Consejo Nacional de Investigaciones Científicas y Técnicas de Argentina (IIBBA-CONICET), Buenos Aires, Argentina 2 Department of Natural Sciences, Bowie State University, Bowie, MD, USA |
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Abstract: | The somatic mutations accumulated in variable and framework regions of antibodies produce structural changes that increase the affinity towards the antigen. This implies conformational and non covalent bonding changes at the paratope, as well as possible quaternary structure changes and rearrangements at the VH-VL interface. The consequences of the affinity maturation on the stability of the Fv domain were studied in a system composed of two closely related antibodies, F10.6.6 and D44.1, which recognize the same hen egg-white lysozyme (HEL) epitope. The mAb F10.6.6 has an affinity constant 700 times higher than D44.1, due to a higher surface complementarity to HEL. The structure of the free form of the Fab F10.6.6 presented here allows a comparative study of the conformational changes produced upon binding to antigen. By means of structural comparison, kinetics and thermodynamics of binding and stability studies on Fab and Fv fragments of both antibodies, we have determined that the affinity maturation process of anti-protein antibodies affects the shape of the combining site and the secondary structure content of the variable domain, stabilizes the VH-VL interaction, and consequently produces an increase of the Fv domain stability, improving the binding to antigen. |
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Keywords: | HEL, hen egg-white lysozyme mAb, monoclonal antibody CDR, complementarity-determining region FR, framework region VH, variable domain of the heavy chain VL, variable domain of the light chain Fab, antigen-binding fraction Fv, variable fraction CH1, first constant domain of the heavy chain CL, constant domain of the light chain rmsd, root-mean-square deviation vdW, van der Waals ANS, 8-anilino-1-naphthalenesulfonic acid |
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