Mutation of Cys-67 alters the thermodynamic stability of the human leukocyte antigen HLA0-B*2705.
The B pocket of the class I major histocompatibility complex-encoded protein HLA-B*2705 has recently been suggested to be responsible for the misfolding of this HLA haplotype and thus to induce susceptibility to autoimmune inflammatory diseases. Four mutants of the B*2705 heavy chain were refolded in the presence of three control peptides. The monitoring of the thermal unfolding of the B*2705-peptide complexes by circular dichroism spectroscopy showed that all heterotrimeric mutants were markedly less stable than the corresponding complexes with the wild-type heavy chain. Among the four heavy chain mutations, the C67S change was investigated for unfolding and peptide binding properties because this position may mediate disulfide pair bridging and alter T-cell recognition of HLA-B*2705. Wild-type heterotrimers completely unfold in a single transition at mild acidic pH whereas increase of the pH to mild basic conditions induce only a partial biphasic unfolding. Cys-67 seems to play a crucial role in controlling the thermodynamic stability of the B*2705-peptide complexes as the C67S mutant unfolds faster and with a single transition, independent of pH. Fluorescence polarization and size exclusion chromatography of unfolding intermediates suggest that the peculiar unfolding of the B*2705 wild-type heavy chain cannot be explained by modified peptide binding properties but more likely by the formation of high molecular weight species.
Department of Applied BioSciences, Swiss Federal Institute of Technology, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
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Reinelt
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Dedier
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Asuni
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Folkers
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Rognan
D003545:Cysteine; D015796:HLA-B27 Antigen / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D006801:Humans; D007962:Leukocytes / Q000276:immunology; D017354:Point Mutation; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D015796:HLA-B27 Antigen; D003545:Cysteine