HLA-B polymorphism affects interactions with multiple endoplasmic reticulum proteins.

Area of Science:

• Immunology
• Molecular Biology
• Protein Biochemistry

Background:

• The assembly of HLA class I molecules with the peptide-loading complex, including TAP (Transporter associated with antigen processing), tapasin, and calreticulin, is crucial for immune surveillance.
• Amino acid residues within the HLA class I peptide-binding groove and surrounding regions can influence these interactions.

Purpose of the Study:

• To investigate how specific amino acid substitutions at position 116 of HLA-B molecules affect their association with TAP and other components of the peptide-loading complex.
• To compare the interaction profiles of a site-directed mutant and natural HLA-B subtypes with varying residues at position 116.

Main Methods:

• Site-directed mutagenesis of HLA-B*0702 to create a Y116D mutant.
• Co-immunoprecipitation assays to assess the association of HLA-B variants with TAP, tapasin, and calreticulin.
• Flow cytometry to evaluate the surface expression of HLA class I molecules.

Main Results:

• The HLA-B*0702 (Y116D) mutant exhibited enhanced association with TAP, tapasin, and calreticulin compared to wild-type HLA-B7.
• Natural HLA-B15 subtypes displayed differential interactions: HLA-B*1510 (Tyrosine at 116) showed strong association, while HLA-B*1518 and HLA-B*1501 (Serine at 116) showed minimal association.
• HLA-B15 subtypes with low intracellular assembly complex association had higher surface expression of the open heavy chain.

Conclusions:

• Position 116 is critical for the association of HLA-B molecules with the TAP complex and associated proteins.
• Amino acid identity at position 116 dictates the strength of interaction with the entire peptide-loading complex.
• Reduced intracellular assembly complex interaction may lead to increased surface stability of HLA class I molecules.