Interaction between heat-shock protein 73 and HLA–DRB1 alleles associated or not with rheumatoid arthritis
Abstract
Objective
HLA–DRB1 alleles whose third hypervariable region contains a QKRAA/QRRAA/RRRAA motif are associated with rheumatoid arthritis (RA) through unknown mechanisms. We previously demonstrated that the QKRAA motif was also expressed on the Escherichia coli 40-kd heat-shock protein (HSP) DnaJ. The QKRAA motif helps DnaJ bind its partner chaperone, the E coli 70-kd HSP DnaK. Furthermore, we observed that in lymphoblastoid cells, Hsp73, the constitutive 70-kd HSP, associates with HLA–DRB1*0401 (an allele with a QKRAA motif) and targets it to lysosomes. In this study, we sought to classify different HLA–DRB1 alleles according to their ability to bind Hsp73.
Methods
To evaluate how well different HLA–DRB1 alleles could bind Hsp73, we developed a quantitative precipitation assay and a direct binding assay.
Results
Quantitative precipitation assay from total cellular proteins and from lysosomal extracts demonstrated that RA-associated HLA–DRB1 alleles bound Hsp73 better than did HLA–DRB1 alleles that were not associated with RA. HLA–DRB1*0401 was the best Hsp73 binder. These findings were confirmed by direct binding assay between purified proteins.
Conclusion
HLA–DRB1*0401 was the best Hsp73 binder among the 8 different HLA–DRB1 alleles that were tested.
