Mouse intracellular immunoglobulin M. Structure and identification of a free thiol group.
Monomeric intracellular mouse immunoglobulin M (hereafter designated IgMs) was purified in milligram quantities from the plasma cells of mouse plasmacytoma MOPC 104E after lysis either in the presence or in the absence of iodoacetate. Peptide ;mapping' analysis of the IgMs after partial reduction and carboxy[(14)C]methylation to label the interchain disulphide bridges showed that the heavy-light bridge and the interheavy bridge present in the Cmu2 region were already formed at lysis. The cysteine residues in the C-terminal region of the heavy chains, which in pentameric IgM form an intersubunit bridge, had free thiol groups at lysis that were reversibly oxidized during isolation in the absence of iodoacetate, probably forming an intrasubunit inter-heavy-chain disulphide bridge. Isoelectric-focusing studies complemented the above findings, showing that all the intracellular IgMs carried free thiol groups that could be carboxymethylated at lysis, and that in non-alkylated preparations these had reversibly oxidized. On the basis of sodium dodecyl sulphate/polyacrylamide-slab-gel electrophoresis intracellular mu-chains had a consistently lower apparent molecular weight than did secreted mu-chains, and the estimated difference could be accounted for by the known difference in carbohydrate content. We present evidence that in a position homologous to that of a complex oligosaccharide in the Cmu2 region of secreted human mu-chains there is a simple oligosaccharide in intracellular mouse mu-chains that becomes complex on secretion. On the basis of the above findings, we present a model for the mouse intracellular IgM subunit and suggest a mechanism for its assembly into secreted IgM pentamers.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D007075:Immunoglobulin M / Q000302:isolation & purification; D007525:Isoelectric Focusing; D051379:Mice; D008961:Models, Structural; D010446:Peptide Fragments / Q000032:analysis; D013438:Sulfhydryl Compounds / Q000032:analysis*
D007075:Immunoglobulin M; D010446:Peptide Fragments; D013438:Sulfhydryl Compounds