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By
From the Center for Cancer Research, Department of Biology, Massachusetts Institute of Technology,
Cambridge, Massachusetts 02139
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 
2-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 2-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.
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