DIFFERENTIAL-T CELL-RECEPTOR PHOTOAFFINITY-LABELING AMONG H-2KD RESTRICTED CYTOTOXIC T-LYMPHOCYTE CLONES SPECIFIC FOR A PHOTOREACTIVE PEPTIDE DERIVATIVE - LABELING OF THE ALPHA-CHAIN CORRELATES WITH J-ALPHA SEGMENT USAGE

Using a direct binding assay based on photoaffinity labeling, we studied the interaction of T cell receptor (TCR) with a K(d)-bound photoreactive peptide derivative on living cells. The K(d)-restricted Plasmodium berghei circumsporozoite (PbCS) peptide 253-260 (YIPSAEKI) was reacted NH2-terminally with biotin and at the TCR contact residue Lys259 with photoreactive iodo, 4-azido salicylic acid (IASA) to make biotin-YIPSAEK(IASA)I. Cytotoxic T lymphocyte (CTL) clones derived from mice immunized with this derivative recognized this conjugate, but not a related one lacking the IASA group nor the parental PbCS peptide. The clones were K(d) restricted. Recognition experiments with variant conjugates, lacking substituents from IASA, revealed a diverse fine specificity pattern and indicated that this group interacted directly with the TCR. The TCR of four clones could be photoaffinity labeled by biotin-YIPSAEK(I-125ASA)I. This labeling was dependent on the conjugates binding to the K(d) molecule and was selective for the TCR alpha (2 clones) or beta chain (1 clone), or was common for both chains (I clone). TCR sequence analysis showed a preferential usage of JalphaTA28 containing alpha chains that were paired with Vbeta1 expressing beta chains. The TCR that were photoaffinity labeled at the alpha chain expressed these Jalpha and Vbeta segments. The tryptophan encoded by the JalphaTA28 segment is rarely found in other Jalpha segments. Moreover, we show that the IASA group interacts preferentially with tryptophan in aqueous solution. We thus propose that for these CTL clones, labeling of the alpha chain occurs via the Jalpha-encoded tryptophan residue.