Folding of HLA-B27 subtypes is determined by the global effect of polymorphic residues and shows incomplete correspondence to ankylosing spondylitis.

Objective: To investigate the maturation and folding of HLA-B27 subtypes and the relationship of these features to ankylosing spondylitis (AS).

Methods: Stable transfectants expressing B27 subtypes and site-directed mutants were used. Maturation/export rates were measured by acquisition of endoglycosidase H resistance. Folding efficiency was estimated from the ratio of unfolded heavy chain to folded heavy chain, which was immunoprecipitated with specific antibodies, in pulse-chase experiments. Association with calnexin was analyzed in coprecipitation experiments. Cytosolic dislocation was estimated by immunoprecipitation of deglycosylated heavy chain after proteasome inhibition. The level of heavy chain expression on unstimulated or interferon-gamma (IFNgamma)-stimulated cells was quantified by Western blotting.

Results: There was no correlation between the export rate and the association of HLA-B27 subtypes with AS. Three of the 4 AS-associated B27 subtypes showed inefficient folding, but B*2707 folded with the same high efficiency as the non-disease-associated subtypes. Some individual mutations that mimicked subtype polymorphism profoundly influenced folding, but in a context-dependent way. The differences in export and folding rates among B27 variants were unrelated to levels of heavy chain expression in the corresponding transfectants, as indicated by the lack of correlation between the two parameters and by heavy chain up-regulation with IFNgamma. Misfolded heavy chain was inefficiently cleared from the endoplasmic reticulum, based on the marginal increase in levels of deglycosylated heavy chain, which resulted from loss of the glycan moiety after cytosolic dislocation, following proteasome inhibition.

Conclusion: HLA-B27 subtype folding is determined by the overall heavy-chain structure, since the effect of a given polymorphism depends on its structural context. Heavy chain misfolding does not explain the association of B*2707 with AS.