Understanding the Energetic Basis for T-cell Receptor Recognition

T-cell receptors (TCRs) are clonotypic, heterodimeric receptors on the surface of T-cells which possess Complementarity Determining Region (CDR) loops similar to immunoglobulins. Their function is to interact with peptides presented on Major-Histocompatiblity Complexes (MHCs) on the surface of most nucleated cells. The TCR of a circulated T-cell (one which has undergone selection in the thymus) binds to an MHC presenting a 'non-self' peptide with a stronger affinity than MHCs presenting 'self' peptides. The TCR must be able to distinguish between these two cases despite the fact that the majority of the potential interface is conserved between the two. To assess to what extent the different portions of the TCR-pMHC interface contribute to binding, we quantify the contributions to binding of the side-chain contacts between the residues at the interface through double-mutant cycles. In a double mutant cycle between two residues, binding experiments are performed between i) the native partners, ii) the two partners with one residue mutated to alanine, iii) the two partners with the other residue mutated to alanine and iv) the two partners with both residues mutated to alanine. The interaction energy between those two residues is defined as native free energy change minus the free energy changes of the two single mutants, plus the free energy change of the double-mutant; ddGint = dG(Xwt-Ywt) - dG(Xz->A,Ywt) - dG(Xwt,Yz->A) + dG(Xz->A,Yz->A). By comparing the interaction energies of the TCR/peptide residue pairs and the TCR/MHC residue pairs, we can approximate the relative contributions of the peptide and the MHC to TCR/pMHC binding. Double-mutant cycles allow us to compare the contributions to affinity from contacts made by the germline CDR 1&2 loops and the variable CDR3 loops between the A6 TCR and the HLA-A2/tax9 pMHC. Our results show a strong favorable interaction energy between CDR3 alpha and the MHC. These data suggest that despite the strong immunological pressures, the energetic profile of TCR recognition may be more malleable or variable than previously thought.