Strategic mutations in the class I major histocompatibility complex HLA-A2 independently affect both peptide binding and T cell receptor recognition.

Mutational studies of T cell receptor (TCR) contact residues on the surface of the human class I major histocompatibility complex (MHC) molecule HLA-A2 have identified a "functional hot spot" that comprises Arg(65) and Lys(66) and is involved in recognition by most peptide-specific HLA-A2-restricted TCRs. Although there is a significant amount of functional data on the effects of mutations at these positions, there is comparatively little biochemical information that could illuminate their mode of action. Here, we have used a combination of fluorescence anisotropy, functional assays, and Biacore binding experiments to examine the effects of mutations at these positions on the peptide-MHC interaction and TCR recognition.

Thermodynamic and kinetic analysis of a peptide-class I MHC interaction highlights the noncovalent nature and conformational dynamics of the class I heterotrimer.

The class I major histocompatibility (MHC) molecule is a heterotrimer composed of a heavy chain, the small subunit beta(2)-microglobulin (beta(2)m), and a peptide. Fluorescence anisotropy has been used to assay the interaction of a labeled peptide with a recombinant, soluble form of the class I MHC HLA-A2. Consistent with earlier work, peptide binding is shown to be a two-step process limited by a conformational rearrangement in the heavy chain/beta(2)m heterodimer.