Selection of functional T cell receptor mutants from a yeast surface-display library

The heterodimeric alpha beta T cell receptor (TCR) for antigen is the key d eterminant of T cell specificity, The structure of the TCR is very similar to that of antibodies, but the engineering of TCRs by directed evolution wi th combinatorial display libraries has not been accomplished to date. Here, we report that yeast surface display of a TCR was achieved only after the mutation of specific variable region residues. These residues are located i n two regions of the TCR at the interface of the alpha- and beta-chains and in the beta-chain framework region that is thought to be in proximity to t he CD3 signal-transduction complex. The mutations are encoded naturally in many antibody variable regions, indicating specific functional differences that have not been appreciated between TCRs and antibodies. The identificat ion of these residues provides an explanation for the inherent difficulties in the display of wild-type TCRs compared with antibodies. Yeast-displayed mutant TCRs bind specifically to the peptide/MHC antigen, enabling enginee ring of soluble T cell receptors as specific T cell antagonists. This strat egy of random mutagenesis followed by selection for surface expression may be of general use in the directed evolution of other eukaryotic proteins th at are refractory to display.