Efficient assembly of recombinant major histocompatibility complex class Imolecules with preformed disulfide bonds

The expression of major histocompatibility class I (MHC-1) crucially depend s upon the binding of appropriate peptides. MHC-1 from natural sources are therefore always preoccupied with peptides complicating their purification and analysis. Here, we present an efficient solution to this problem. Recom binant MHC-1 heavy chains were produced in Escherichia coli and subsequentl y purified under denaturing conditions. In contrast to common practice, the molecules were not reduced during the purification. The oxidized MHC-I hea vy chain isoforms were highly active with respect to peptide binding. This suggests that de novo folding of denatured MHC-I molecules proceed efficien tly if directed by preformed disulfide bond(s). Importantly, these molecule s express serological epitopes and stain specific T cells; and they bind pe ptides specifically. Several denatured MHC-1 heavy chains were analyzed and shown to be of a quality, which allowed quantitative analysis of peptide b inding. The analysis of the specificity of the several hundred human MHC ha plotypes, should benefit considerably from the availability of pre-oxidized recombinant MHC-I.