Glycosylation of Pea Cotyledon Membranes

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Abstract
Pea cotyledons were injected with d-[14C]mannose or d-[14C]-glucosamine and incubated for 1 to 1.5 hours. Cotyledons were homogenized and subcellular fractions were isolated by differential centrifugation followed by linear sucrose density gradient centrifugation. Radioactivity that was precipitated by trichloroacetic acid was associated most extensively with rough endoplasmic reticulum, Golgi membranes, a membrane with a density of 1.14 grams per cubic centimeter (possibly plasma membrane) and an unidentified subcellular component with a density of 1.22 grams per cubic centimeter. Lower levels of incorporation were observed in protein bodies and mitochondria. Isolated membrane fractions were lipid-extracted to determine which components of the membrane contained the label. Rough endoplasmic reticulum contained the most extensively labeled lipids which had similar properties to the lipid intermediates thought to be involved in glycoprotein assembly. The lipid free residues of the various membrane fractions contained radioactivity that was released by protease treatment. Acid hydrolysis of the residues indicated that most of the radioactivity was associated with mannose or glucosamine. It appears that various subcellular components of the pea cotyledon possess glycoproteins that contain mannose and glucosamine.