Oligosaccharide processing at individual glycosylation sites on MOPC 104E immunoglobulin M. Differences in α1,2-linked mannose processing

Processing of the asparagine-linked oligosaccharides at the known glycosylation sites on the μ-chain of IgM secreted by MOPC 104E murine plasmatocytoma cells was investigated. Oligosaccharides present on intracellular μ-chain precursors were of the high mannose type, remaining susceptible to endo-β-N-acetylglucosaminidase H. However, only 26% of the radioactivity was released from [³H]mannose-labeled secreted IgM glycopeptides, consistent with the presence of high mannose-type and complex-type oligosaccharides on the mature μ-chain. [³H]mannose-labeled cyanogen bromide glycopeptides derived from μ-chains of secreted IgM were isolated and analyzed to identify the glycopeptide containing the high mannose-type oligosaccharide from those containing complex-type structures. [³H]Mannose-labeled intracellular μ-chain cyanogen bromide glycopeptides corresponding to those from secreted IgM were isolated also, and the time courses of oligosaccharide processing at the individual glycosylation sites were determined. The major oligosaccharides on all intracellular μ-chain glycopeptides after 20 min of pulse labeling with [³H]mannose were identified as Man₈GlcNAc₂, Man₉GlcNAc₂, and Glc₁Man₉GlcNAc₂. Processing of the oligosaccharide destined to become the high mannose-type structure on the mature protein was rapid. After 30 min of chase incubation the predominant structures of this oligosaccharide were Man₅GlcNAc₂ and Man₆GlcNAc₂ which were also identified on the high mannose-type oligosaccharide of the secreted μ-chain. In contrast, processing of oligosaccharides destined to become complex type was considerably slower. Even after 180 min of chase incubation, Man₇GlcNAc₂ and Man₈GlcNAc₂ were the predominant structures at some of these glycosylation sites. The isomeric structures of Man₈GlcNAc₂ obtained from all of the glycosylation sites were identical. Thus, the different rates of processing were not the result of a different sequence of α1,2-mannose removal.