title: Nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase is phosphorylated in wheat endosperm at serine-404 by an SNF1-related protein kinase allosterically inhibited by ribose-5-phosphate.
abstract: Nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase (np-Ga3PDHase) is a cytosolic unconventional glycolytic enzyme of plant cells regulated by phosphorylation in heterotrophic tissues. After interaction with 14-3-3 proteins, the phosphorylated enzyme becomes less active and more sensitive to regulation by adenylates and inorganic pyrophosphate. Here, we acknowledge that in wheat (Triticum aestivum), np-Ga3PDHase is specifically phosphorylated by the SnRK (SNF1-related) protein kinase family. Interestingly, only the kinase present in heterotrophic tissues (endosperm and shoots, but not in leaves) was found active. The specific SnRK partially purified from endosperm exhibited a requirement for Mg(2+) or Mn(2+) (being Ca(2+) independent), having a molecular mass of approximately 200 kD. The kinase also phosphorylated standard peptides SAMS, AMARA, and SP46, as well as endogenous sucrose synthase, results suggesting that it could be a member of the SnRK1 subfamily. Concurrently, the partially purified wheat SnRK was recognized by antibodies raised against a peptide conserved between SnRK1s from sorghum (Sorghum bicolor) and maize (Zea mays) developing seeds. The wheat kinase was allosterically inhibited by ribose-5-phosphate and, to a lesser extent, by fructose-1,6-bisphosphate and 3-phosphoglycerate, while glucose-6-phosphate (the main effector of spinach [Spinacia oleracea] leaves, SnRK1) and trehalose-6-phosphate produced little or no effect. Results support a distinctive allosteric regulation of SnRK1 present in photosynthetic or heterotrophic plant tissues. After in silico analysis, we constructed two np-Ga3PDHase mutants, S404A and S447A, identifying serine-404 as the target of phosphorylation. Results suggest that both np-Ga3PDHase and the specific kinase could be under control, critically affecting the metabolic scenario involving carbohydrates and reducing power partition and storage in heterotrophic plant cells.
authors.affiliation: Instituto de Agrobiotecnología del Litoral, (Consejo Nacional de Investigaciones Científicas y Técnicas-Universidad Nacional del Litoral), Facultad de Bioquímica y Ciencias Biológicas, Paraje El Pozo, S3000ZAA Santa Fe, Argentina.
authors.forename: Claudia Vanesa
authors.initials: CV
authors.lastname: Piattoni
authors.forename: Diego Martín
authors.initials: DM
authors.lastname: Bustos
authors.forename: Sergio Adrián
authors.initials: SA
authors.lastname: Guerrero
authors.forename: Alberto Álvaro
authors.initials: AÁ
authors.lastname: Iglesias
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects; D000595:Amino Acid Sequence; D002413:Cations, Divalent / Q000494:pharmacology; D056625:Endosperm / Q000187:drug effects / Q000201:enzymology*; D005635:Fructosediphosphates / Q000494:pharmacology; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D005988:Glyceric Acids / Q000494:pharmacology; D007700:Kinetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000187:drug effects; D010455:Peptides / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D012267:Ribosemonophosphates / Q000494:pharmacology*; D016415:Sequence Alignment; D014908:Triticum / Q000187:drug effects / Q000201:enzymology*
chemical_list: D002413:Cations, Divalent; D005635:Fructosediphosphates; D005988:Glyceric Acids; D010455:Peptides; D012267:Ribosemonophosphates; D010768:Phosphoserine; C031626:ribose-5-phosphate; C005156:3-phosphoglycerate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; C071570:SNF1-related protein kinases; D017346:Protein Serine-Threonine Kinases; C029063:fructose-1,6-diphosphate