An extensive region of an MHC class I alpha 2 domain loop influences interaction with the assembly complex

Presentation of antigenic peptides to CTLs at the cell surface first requir es assembly of R-IHC class I with peptide and beta(2)-microglobulin in the endoplasmic reticulum, This process involves an assembly complex of several proteins, including TAP, tapasin, and calreticulin, all of which associate specifically with the beta(2)-microglobulin-assembled, open form of the cl ass I heavy chain. To better comprehend at a molecular level the regulation of class I assembly, we have assessed the influence of multiple individual amino acid substitutions in the MHC class I alpha 2 domain on interaction with TAP, tapasin, and calreticulin. In this report, we present evidence in dicating that many residues surrounding position 134 in H-2L(d) influence i nteraction with assembly complex components. Most mutations decreased assoc iation, but one (L(d)K131D) strongly increased it. The L-d mutants, with th e exception of L(d)K131D, exhibited characteristics suggesting suboptimal i ntracellular peptide loading, similar to the phenotype of Ld expressed in a tapasin-deficient cell line. Notably, K131D was less peptide inducible tha n wild-type L-d, which is consistent with its unusually strong association with the endoplasmic reticulum assembly complex.