Empty and Peptide-Loaded Class II Major Histocompatibility Complex Proteins Produced by Expression inEscherichia coliand Foldingin Vitro

The human class II major histocompatibility complex protein HLA–DR1 has been expressed inEscherichia colias denatured α and β subunits and foldedin vitroto form the native structure. DR1 folding yields are 30–50% in the presence or absence of tight-binding antigenic peptides. The protein produced in this manner is soluble and monomeric with the expected apparent molecular weight. It reacts with conformation-sensitive anti-DR antibodies and exhibits peptide-dependent resistance to SDS-induced chain dissociation and to proteolysis as does the native protein. The observed peptide specificity and dissociation kinetics are similar to those of native DR produced in B-cells and finally the protein exhibits circular dichroism spectra and cooperative thermal denaturation as expected for a folded protein. We conclude that the recombinant DR1 has adopted the native fold. We have folded DR1 in the absence of peptide and isolated a soluble, peptide-free αβ-heterodimer. The empty DR1 can bind antigenic peptide but exhibits altered far UV-circular dichroism and thermal denaturation relative to the peptide-bound form.