Sodium dodecyl sulfate stability of HLA-DR1 complexes correlates with burial of hydrophobic residues in pocket 1

Certain class II MHC-peptide complexes are resistant to SDS-induced dissoci ation, This property, which has been used as an in vivo as well as an in vi tro peptide binding assay, is not understood at the molecular level. Here w e have investigated the mechanistic basis of SDS stability of HLA-DR1 compl exes by using a biosensor-based assay and SDS-PAGE with a combination of wi ld-type and mutant HLA-DR1 and variants of hemagglutinin peptide HA(306-318 ). Experiments with wild-type DR1 along with previously published results e stablish that the SDS-stable complexes are formed only when the hydrophobic pocket 1 (P1) is occupied by a bulky aromatic (Trp, Phe, Tyr) or an alipha tic residue (Met, Ile, Val, Leu). To further explore whether the SDS sensit ivity is primarily due to the exposed hydrophobic regions, we mutated resid ue beta Gly(86) at the bottom of P1 to tyrosine, presumably reducing the de pth of the pocket and the exposure of hydrophobic residues and increasing t he contacts between subunits. In direct contrast to wild-type DR1, the pept ide-free mutant DR1 exists as an alpha/beta heterodimer in SDS, Moreover, t he presence of a smaller hydrophobic residue, such as alanine, as P1 anchor with no contribution from any other anchor is sufficient to enhance the SD S stability of the mutant complexes, demonstrating that the basis of SDS re sistance may be localized to PI interactions. The good correlation between SDS sensitivity and the exposure of hydrophobic residues provides a biochem ical rationale for the use of this assay to investigate the maturation of c lass II molecules and the longevity of the complexes.