Unassembled (free) heavy chains appear during two stages of the class I MHC molecule's existence: immediately after translation but before assembly with peptide and β2-microglobulin, and later, upon disintegration of the heterotrimeric complex. To characterize the structures of folding and degradation intermediates of the class I heavy chain, three monoclonal antibodies have been produced that recognize epitopes along the H-2Kb heavy chain which are obscured upon proper folding and subsequent assembly with β2-microglobulin (KU1: residues 49-54; KU2: residues 23-30; KU4: residues 193-198). The Kb heavy chain is inserted into the lumen of the endoplasmic reticulum in an unfolded state reactive with KU1, KU2, and KU4. Shortly after completion of the polypeptide chain, reactivity with KU1, KU2 and KU4 is lost synchronously, suggesting that folding of the class I heavy chain is a rapid, cooperative process. Perturbation of the folding environment in intact cells with the reducing agent dithiothreitol or the trimming glucosidase inhibitor N-7-oxadecyl-deoxynojirimycin prolongs the presence of mAb-reactive Kb heavy chains. At the cell surface, a pool of free Kb heavy chains appears after 60–120 min of chase, whose subsequent degradation, but not their initial appearance, is impaired in the presence of concanamycin B, an inhibitor of vacuolar acidification. Thus, free heavy chains that arise at the cell surface are destroyed after internalization.
Targeting cell surface β2-microglobulin by pentameric IgM antibodies