Folding and Assembly of a Human MHC Class II Molecule in a Cell-Free System

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Abstract
The assembly of a human major histocompatibility complex (MHC) class II molecule was investigated in a cell-free system capable of the synthesis, sequestration, and processing of the protein chains. As assessed by the conformation-sensitive monoclonal antibody L243, the formation of HLA-DR α/β heterodimer required cotranslation of α and β mRNA in the presence of both oxidized glutathione and canine pancreas endoplasmic reticulum (ER) vesicles. The assembly of α/β dimer could also be initiated by the post-translational addition of oxidized glutathione. Using the post-translational assay system, we investigated the effect of the depletion of ER lumenal content proteins on the folding and assembly of the MHC class II chains. Both the rate and extent of folding of α chain and β chain and the post-translational assembly of α/β dimer is greatly reduced in the depleted ER vesicles. Conversely, the extent of aggregate formation is increased. Upon reconstitution of the depleted ER vesicles with lumenal proteins, the folding of α chain is accelerated and the assembly of α/β dimer is increased.

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