A soluble, single‐chain K^d molecule produced by yeast selects a peptide repertoire indistinguishable from that of cell‐surface‐associated K^d

Peptide binding to a soluble, single‐chain K^d protein produced by the yeast strain Kluyveromyces lactis, and to K^d molecules on K^d‐expressing cells (P815) was studied using radiolabeled K^d‐restricted peptides. The stability of the peptide‐K^d complexes formed was monitored in the absence and presence of unlabeled competitor peptides. Radioiodination of the Tyr anchor residue in position 2 of the peptide interferes with binding. A K^d‐biased peptide library and a modified antigenic peptide in which a second Tyr was added in positions 6 and 8, respectively, were therefore used to assay binding. Recombinant and cell‐associated K^d molecules are very similar in the following respects: the ease with which the proteins can be loaded with labeled peptide; the spectrum of peptides selected from a peptide library; the stability of the labeled peptide‐K^d complex formed; and the ability to partially dissociate the class 1‐peptide complex with exogenous, unlabeled peptides. These results imply that measurements of peptide binding to soluble K^d molecules are a reliable indicator of the peptide‐binding properties of K^d proteins on living cells. The large quantities of soluble recombinant K^d protein currently available represent an invaluable tool not only for dissecting the molecular mechanisms of antigen presentation but also for vaccinations and the design of T cell‐specific toxins.