title: Demonstration of glucose-6-phosphate hydrogen 5 enrichment from deuterated water by transaldolase-mediated exchange alone.
abstract: OBJECTIVE
  Enrichment of glucose position 5 (H5) from deuterated water ((2)H2O) is widely used for quantifying gluconeogenesis. Exchanges of hexose and triose phosphates mediated by transaldolase have been postulated to enrich H5 independently of gluconeogenesis, but to date this mechanism has not been proven. We determined the enrichment of glucose-6-phosphate (G6P), the immediate precursor of endogenously produced glucose, from (2)H2O in erythrocyte hemolysate preparations. Here, transaldolase exchange is active but gluconeogenesis is absent.
  
  
  METHODS
  Hemolysates were prepared from human erythrocytes and incubated with a buffer containing 5% [U-(13)C]G6P, unlabeled fructose 1,6-bisphosphate, and 10% (2)H2O. G6P (2)H-enrichment and (13)C-isotopomer distributions were analyzed by (2)H and (13)C NMR following derivatization to monoacetone glucose.
  
  
  RESULTS
  (2)H NMR analysis revealed high (2)H-enrichment of G6P hydrogens 2, 4, and 5; low enrichment of hydrogen 3, and residual enrichments of hydrogens 1, 6R, and 6S. (13)C NMR isotopomer analysis revealed that [U-(13)C]G6P was converted to [1,2,3-(13)C3]G6P, a predicted product of transaldolase-mediated exchange, as well as [1,2-(13)C2]G6P and [3-(13)C]G6P, predicted products of combined transaldolase and transketolase exchanges.
  
  
  CONCLUSIONS
  Hydrogen 5 of G6P was enriched from (2)H2O through exchanges mediated by transaldolase. These studies prove that G6P can be enriched in hydrogen 5 by (2)H2O independently of gluconeogenesis.
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.
authors.forename: Margarida
authors.initials: M
authors.lastname: Coelho
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.
authors.forename: Paula
authors.initials: P
authors.lastname: Valente-Silva
authors.affiliation: Department of Pediatrics, Nutrition and Metabolic Disease, The Children's Memorial Health Institute, Warsaw, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Tylki-Szymanska
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.
authors.forename: Tiago
authors.initials: T
authors.lastname: Henriques
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.
authors.forename: Cristina
authors.initials: C
authors.lastname: Barosa
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.
authors.forename: Filipa
authors.initials: F
authors.lastname: Carvalho
authors.affiliation: Center for Neurosciences and Cell Biology, University of Coimbra, Portugal.|APDP-Portuguese Diabetes Association, Lisbon, Portugal.
authors.forename: John G
authors.initials: JG
authors.lastname: Jones
mesh_terms: D017666:Deuterium Oxide / Q000737:chemistry* / Q000378:metabolism*; D004912:Erythrocytes / Q000737:chemistry / Q000378:metabolism; D005943:Gluconeogenesis; D019298:Glucose-6-Phosphate / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D014153:Transaldolase / Q000378:metabolism*
chemical_list: D019298:Glucose-6-Phosphate; D014153:Transaldolase; D017666:Deuterium Oxide
keywords: 13C-NMR isotopomer analysis; deuterated water; gluconeogenesis; pentose phosphate pathway; transaldolase exchange