Evidence For The Glycoprotein Nature Of Retina Glycogen

Incubation of a bovine retina membrane preparation with micromolar amounts of UDP-[14C]glucose resulted in the incorporation of [14C]glucose into endogenous (1→4)-α-glucan, insoluble in trichloroacetic acid, and acidsoluble ethanol-insoluble glycogen. 1 The trichloroacetic-acid-insoluble glucan fraction of retina migrated in 2.6–3% acrylamide gels when subjected to sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS-PAGE) and was rendered acidsoluble by digestion with pronase. 2 The solubility of the acid-insoluble glucan in acidified organic solvent was different from that of amylose or glycogen and similar to membrane proteins and glycoproteins. 3 The glycogen fraction of retina contained 1.5–2.0 μ protein/100 μ glucose. When this fraction was analyzed by SDS-PAGE only one band, which moved near the top of 3% acrylamide gels, was stained with periodic acid Schiff reagent and Coomassie blue. 4 The protein nature of the Coomassie-blue-stainable material was demonstrated by iodination of the glycogen fraction with [131I]iodide and identification of labeled monoiodotyrosine and diiodotyrosine. The bulk of the label comigrated with carbohydrate near the top of gels in SDS-PAGE and treatmet with α-amylse decreased the molecular size of both labeled and stainable material. 5 Physical dissociative conditions (7.5 M urea/0.83% mercaptoethanol) and the following chemical tratments failed to dissociate the iodinated protein from glycogen: (a) 0.1 M NaOH/0.1 M NaBH4 at room temperature for 24h; (b) 1 M HCl in methanol at 50°C for 10 min; (c) trifluoroacetic acid at 50°C for 6 min. 6 131I-labeled glycogenpeptide was isolated after 131I-labeled protein-bound glycogen had been subjected to digestion with papain/pronase and passed through a Sepharose column. 7 The results suggest that at least part of glycogen in bovine retina is firmly combined to protein as a single proteoglycogen molecule. Furthermore some of the proteoglycogen might be present as a trichloroacetic-acid-precipitable proteoglucan owing to its lower glucose content.