Studies will be undertaken with model antibodies to disect the contribution of different portions of the antibody variable regions to antigen binding. One series of experiments will determine the structural requirements for high affinity binding by a stepwise conversion of the amino acid sequence of a lower affinity mouse antibody directed against azophenylarsonate (Ars) into the sequence of a higher affinity one. Another series of experiments will involve interchanging portions of the variable regions of a mouse anti-Ars antibody with those of a mouse anti-Alpha(1-6) dextran antibody. This will include testing the hypothesis that the hypervariable regions alone carry the antibody specificity and will retain it when inserted into another framework. The above experiments will be accomplished using site directed in vitro mutagenesis of cloned antibody genes, and introduction of these genes into appropriate myeloma or hybridoma cell lines. The mutant antibodies produced will be tested for antigen binding specificity and affinity and for idiotypic expression using standard immunological methods. These studies will increase our understanding of the structural requirements for antibody specificity and affinity, and of the structure - function relationship and folding of proteins in general. They will help develop the knowledge to design tailor - made antibodies for research and medical purposes.
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