Structure of the polysaccharide formed by incubating glycogen with D-[14C]glucose in the presence of the glycogen debranching enzyme [amylo-(1 linked to 6)-glucosidase-4-alpha-glucanotransferase].
[14C]Glycogen has been synthetized in vitro by incubating D-[14C]glucose with rabbit-liver glycogen in the presence of a pure preparation of the glycogen debranching enzyme [amylo-(1 linked to 6)-glucosidase-4-alpha-glucanotransferase]. The course of the reaction has been monitored and 14C-products isolated after 30 min and 5 h. The distribution of D-[14C]glucose groups in the polysaccharides has been determined by debranching the molecules with a crystalline isoamylase from Pseudomonas. The quantities of unlabeled and 14C-linear unit chains containing D-[14C]glucose at their reducing ends have been determined by paper chromatography followed by enzymic degradation and analysis. In the 30-min product, between 65 and 85% of the D-[14C]glucose groups were covered by unlabeled groups because of transferase action. In the 5-h product, the extent of covering approached 100%. Extensive redistribution of unlabeled groups also was found to have occurred, even in the early stages of the reaction. It is concluded that the D-[14C]glucose incorporation assay for amylo-(1 linked to 6)-glucosidase, as ordinarily carried out, is probably not specific just for the hydrolytic action of this enzyme, but that it depends indirectly on its transferase activity as well.
D000818:Animals; D002250:Carbon Radioisotopes; D005947:Glucose / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism*; D006004:Glycogen Debranching Enzyme System / Q000378:metabolism*; D007517:Isoamylase; D008112:Liver Glycogen / Q000378:metabolism*; D009132:Muscles / Q000201:enzymology; D011134:Polysaccharides / Q000032:analysis; D011817:Rabbits; D013997:Time Factors
D002250:Carbon Radioisotopes; D006004:Glycogen Debranching Enzyme System; D008112:Liver Glycogen; D011134:Polysaccharides; D005964:Glucosyltransferases; D007517:Isoamylase; D005947:Glucose