Structural analysis of the asparagine-linked oligosaccharides of rat haptoglobin metabolically labeled in a hepatocyte culture system

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Abstract
We analyzed the asparagine‐linked oligosaccharide chains of rat haptoglobin which were synthesized and secreted by hepatocytes in primary culture. When the cells were incubated with either [3H]mannose, [3H]galactose, or[3H]fucose, all the radioactive precursors were incorporated into the β subunit of haptoglobin. [3H]Mannose‐labeled haptoglobin was purified from the culture medium by immunoaffinity chromatography, and [3H]oligosaccharides were prepared by strong alkali‐borohydride treatment. The oligosaccharides obtained were analyzed by anion‐exchange high‐performance liquid chromatography, concanavalin‐A–Sepharose chromatography and Bio‐Gel P‐4 chromatography before and after sequential exoglycosidase digestions. The oligosaccharides labeled with [3H]fucose or [3H]galactose were also characterized by the above methods. The results indicate that rat haptoglobin contains two complex‐type oligosaccharide chains in each β subunit; one with a possible structure of (± NeuAc→Galβ→GlcNAcβ→) 3(Manα→)2Manβ→GlcNAc→(±Fucα→)GlcNAc and the other with (±NeuAc→Galβ→GlcNAcβ→Manα→)2Manβ→Glc NAc→(±Fucα→)GlcNAc.

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