Antibodies and enzymes are bound to specific substrates, and are therefore called sensing functional protein. A technique of changing their specificity according to the need was required to apply proteins to biosensors or catalysts. Consquently, we proposed two-step strategies for the efficient conversion of the function of proteins using the directed evolution method. For a model case, the specificity for 11-deoxycortisol of a monoclonal antibody was changed to the specificity for cortisol (CS), whose structures were almost identical, with the absence or presence of a hydroxy group at the 11th carbon of the steroid ring.

Antigen specificity of an antibody was determined primarily by the sequences of the antigen binding pocket. As the first step, mutations were introduced at 14 amino acid that seemed to form the binding pocket. A clone, DcC16, was isolated from the resultant library which shown to have CS-binding activity. As the second step, mutations were introduced randomly into the region around the pocket of DcC16 clone to fine tune the binding pocket, and CS-specific mutants were selected. Structural models, constructed by computer simulation, indicated the probable molecular basis for these changes in specificity.
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