Intracellular transport of class I HLA molecules is affected by polymorphic residues in the binding groove.
Intracellular transport of class I MHC complexes is dependent on assembly of class I heavy chains with beta 2-microglobulin (beta 2m) and peptides. This suggests that amino acid residues of individual class I molecules which are important for their stability and transport are likely to include those which contribute to binding of a majority of the cleft-associated peptides. To identify such critical residues, substitutions at polymorphic positions within the peptide binding cleft were introduced into a mutant HLA-A*0201 molecule bearing an additional gly > lys substitution at position 242 (242K). The 242K mutation weakens association of the HLA-A*201 heavy chain with beta 2m and was used to enhance potential effects of substitutions in the peptide binding groove on class I stability. Critical in choosing which binding cleft positions to mutate was the observation that HLA-A*6801 was less sensitive to the effects of 242K mutation than HLA-A*0201 and A*6901. This suggested that one or more of the six residues in the alpha 2 domain differing between HLA-A*6901 and A*6801 were likely to affect class I complex stability. Positions 95, 97, 107, 114, 116, and 156 in either 242K or wild-type HLA-A*0201 molecules were therefore each converted to those residues found in HLA-A*6801. One of the second-site substitutions, arg > met at position 97, increased stability and restored surface expression of the 242K molecule. Five other substitutions either had no additional effect or further impaired 242K stability. Substitution of his > arg at position 114 blocked surface expression of both 242K and wild-type HLA-A*0201 molecules. These results demonstrate that polymorphic residues in the binding cleft influence the stability of class I complexes, and suggest that position 97 plays a critical role in stabilizing class I molecules for transport.
Salter
R D
RD
Department of Pathology, University of Pittsburgh School of Medicine, PA 15261.
D001665:Binding Sites / Q000235:genetics; D001693:Biological Transport, Active / Q000235:genetics; D002460:Cell Line; D004355:Drug Stability; D015870:Gene Expression; D015234:HLA-A Antigens / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D011110:Polymorphism, Genetic; D014162:Transfection
D015234:HLA-A Antigens