Activation of glycogen phosphorylase with 5-aminoimidazole-4-carboxamide riboside (AICAR). Assessment of glycogen as a precursor of mannosyl residues in glycoconjugates.
The experimental evaluation of the contribution of glycogen phosphorylase (GP) to biochemical pathways is limited to methods that raise cAMP, activating the cAMP-dependent protein kinase/phosphorylase kinase/GP cascade. Such methods convert the unphosphorylated form, "GPb," which catalyzes glycogenolysis only in the presence of appropriate allosteric activators such as AMP, to the phosphorylated, constitutively activated form, "GPa." However, activation of GP in this way is indirect, requires a functional cAMP kinase cascade, and is complicated by other actions of cAMP. Here, we demonstrate a strategy for the experimental manipulation of GP in intact dermal fibroblasts, involving activation by the membrane-permeable adenosine analog 5-aminoimidazole-4-carboxamide riboside (AICAR) and inhibition by caffeine and Pfizer compound CP-91149, which bind to GP at distinct sites. Potential complications because of activation of AMP-activated protein kinase by AICAR were assessed with metformin, which activates this kinase but does not activate GP. Using this strategy, we show that glycogen can be a significant and regulatable precursor of mannosyl units in lipid-linked oligosaccharides and glycoproteins.
Shang
Jie
J
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390-9041, USA.
Lehrman
Mark A
MA
D000577:Amides / Q000494:pharmacology; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology*; D001665:Binding Sites; D002110:Caffeine / Q000494:pharmacology; D002384:Catalysis; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D000242:Cyclic AMP / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D004789:Enzyme Activation; D005347:Fibroblasts / Q000378:metabolism; D006003:Glycogen / Q000378:metabolism*; D024981:Glycogen Phosphorylase / Q000378:metabolism*; D006801:Humans; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology; D050356:Lipid Metabolism; D008358:Mannose / Q000737:chemistry / Q000378:metabolism*; D008687:Metformin / Q000494:pharmacology; D009844:Oligosaccharides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D012265:Ribonucleotides / Q000494:pharmacology*; D012867:Skin / Q000166:cytology; D013997:Time Factors
D000577:Amides; C110844:CP 91149; D007004:Hypoglycemic Agents; D007211:Indoles; D009844:Oligosaccharides; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; D002110:Caffeine; D006003:Glycogen; D008687:Metformin; D000242:Cyclic AMP; D024981:Glycogen Phosphorylase; C031143:AICA ribonucleotide; D006152:Cyclic GMP; D008358:Mannose