Calnexin influences folding of human class I histocompatibility proteins but not their assembly with beta 2-microglobulin.
Class I major histocompatibility complex heavy chains bind to calnexin before associating with beta 2-microglobulin (beta 2m) and peptides. Calnexin has been shown to retain in the endoplasmic reticulum those class I heavy chains which have not assembled properly and, thus, to serve as a quality control mechanism. In addition, calnexin may direct the folding of class I subunits or their subsequent assembly. We asked whether calnexin plays a role in the initial folding of HLA-B*0702 heavy chains by assessing disulfide bond formation in vivo. Our results show that class I heavy chains form intrachain disulfide bonds very soon after translation, and that calnexin is bound to both reduced and oxidized forms during this process. When a cell-permeable reducing agent, dithiothreitol, was added to cells, disulfide bond formation in newly synthesized heavy chains was substantially blocked, as was their association with calnexin. The reducing agent appeared to affect calnexin directly, since binding was similarly abolished to a subset of proteins which do not contain internal disulfide bonds. Addition of the glucosidase inhibitor castanospermine to cells, shown previously to disrupt calnexin binding to ligands, slowed formation of disulfide bonds but did not decrease the amount of assembled heavy chain-beta 2m complexes that formed. Our data suggest that calnexin can promote disulfide bond formation in class I heavy chains but does not directly facilitate subsequent binding to beta 2m.
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Department of Pathology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Salter
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D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D037281:Calnexin; D002460:Cell Line; D004220:Disulfides / Q000737:chemistry; D015235:HLA-B Antigens / Q000378:metabolism*; D006801:Humans; D010084:Oxidation-Reduction; D017510:Protein Folding; D001613:beta 2-Microglobulin / Q000378:metabolism*
D002135:Calcium-Binding Proteins; D004220:Disulfides; D015235:HLA-B Antigens; D001613:beta 2-Microglobulin; D037281:Calnexin