title: Rate-determining steps in the biosynthesis of glycogen in COS cells.
abstract: Consistent with previous results, overexpression of rabbit skeletal muscle glycogen synthase in COS cells did not lead to overaccumulation of glycogen unless activating Ser-->Ala mutations were present at key regulatory phosphorylation sites 2 (Ser7) and 3a (Ser644) in the enzyme. In addition, we found that expression of glycogenin, glycogen branching enzyme, or UDP-glucose pyrophosphorylase alone in COS cells had no effect on the glycogen level. However, coexpression of the hyperactive 2,3a glycogen synthase mutant with either glycogenin or UDP-glucose pyrophosphorylase led to higher glycogen accumulation than that obtained from the expression of glycogen synthase alone. Coexpression of glycogenin with the 2,3a mutant of glycogen synthase led to the appearance of glycogenin with a lower molecular weight suggestive of reduced glucosylation. Increased glycogen synthesis may lead to competition between glycogenin and glycogen synthase for their common substrate UDP-glucose. In summary, we conclude that (i) glycogen synthase is a primary rate-limiting enzyme of glycogen biosynthesis in COS cells, (ii) that phosphorylation of glycogen synthase is regulatory for glycogen accumulation, and (iii) once glycogen synthase is activated, the reaction mediated by UDP-glucose pyrophosphorylase can become rate-determining.
authors.affiliation: Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, 46202-5122, USA.
authors.forename: A V
authors.initials: AV
authors.lastname: Skurat
authors.forename: H L
authors.initials: HL
authors.lastname: Peng
authors.forename: H Y
authors.initials: HY
authors.lastname: Chang
authors.forename: J F
authors.initials: JF
authors.lastname: Cannon
authors.forename: P J
authors.initials: PJ
authors.lastname: Roach
mesh_terms: D015061:1,4-alpha-Glucan Branching Enzyme / Q000096:biosynthesis / Q000378:metabolism; D000409:Alanine; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D005822:Genetic Vectors; D005964:Glucosyltransferases; D006003:Glycogen / Q000096:biosynthesis*; D006006:Glycogen Synthase / Q000096:biosynthesis / Q000378:metabolism*; D006023:Glycoproteins / Q000096:biosynthesis / Q000378:metabolism; D007700:Kinetics; D018482:Muscle, Skeletal / Q000201:enzymology*; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012694:Serine; D013997:Time Factors; D014162:Transfection; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase / Q000096:biosynthesis / Q000378:metabolism
chemical_list: D006023:Glycoproteins; D011994:Recombinant Proteins; C051887:glycogenin; D012694:Serine; D006003:Glycogen; D005964:Glucosyltransferases; D006006:Glycogen Synthase; D015061:1,4-alpha-Glucan Branching Enzyme; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase; D000409:Alanine