The TCR from the alloreactive T cell clone 2C recognizes the QL9 (QLSPFPFDL) / Ld complex with an affinity of 107 M-1 (Sykulev el al. PNAS 91:1487, 1994). In order to examine the contact residues involved in this interaction, we have recently expressed the TCR as a soluble single-chain Vβ8.2Vα3.1 in E. coli. This scTCR bound specifically to cell surface QL9/Ld complexes. Using this system, we analyzed the role of a specific QL9 residue (F5), predicted to be involved in TCR binding (Al-Ramadi et. al. J. Immunol. 155:662, 1995). The Phe at position 5 in QL9 was changed to various other amino acids. Relative binding affinities of these peptide/Ld complexes for the 2C TCR were determined and only a Tyr substitution at position 5 is able to retain high affinity for the TCR. Unexpectedly, position 5 variants not only had major effects on TCR binding, but they dramatically affected (100-fold) binding to Ld. These results may suggest that the TCR recognizes an Ld conformational determinant that is altered by interactions with the residue at position 5 of QL9. To begin to examine the TCR residues that interact with QL9/Ld, we have expressed a family of 2C scTCRs which contain alanine point mutations in the CDR and hypervariable 4 (HV4) loops of both the Vα and Vβ chains. Preliminary evidence indicated that Vβ HV4 alanine point mutants retain the ability to bind QL9/Ld. The ability of additional scTCR mutants to bind both QL9/Ld and various monoclonal anti-TCR antibodies is under investigation.
|Original language||English (US)|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Molecular Biology