title: Diphosphosinositol polyphosphates and energy metabolism: assay for ATP/ADP ratio.
abstract: Several inositol compounds undergo rapid cycles of phosphorylation and dephosphorylation. These cycles are dependent on ATP and energy metabolism. Therefore, interfering with the cellular energy metabolism can change the concentration of rapidly turning over inositols. Many pharmacological inhibitors, apart from their intended action, also affect the energy metabolism of the cells and lower ATP. This can unspecifically influence rapidly turning over inositol phosphates. Thus, the ATP concentration should be checked when reduced inositol phosphates are observed after application of pharmacological inhibitors. A luminescence-based assay for the measurement of ATP and ADP is described. ATP is measured luminometrically using firefly luciferase. Detection of ADP is performed in a two-step enzymatic procedure: (1) The sample ATP is degraded to AMP and (2) ADP is phosphorylated to ATP, which can then be measured luminometrically. This method gives a better signal-to-noise ratio than other methods that do not degrade the sample ATP, but convert ADP directly to ATP and then measure the sum of ATP plus ADP.
authors.affiliation: The Rolf Luft Research Center for Diabetes and Endocrinology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Andreas
authors.initials: A
authors.lastname: Nagel
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Barker
authors.forename: Per-Olof
authors.initials: PO
authors.lastname: Berggren
authors.forename: Christopher
authors.initials: C
authors.lastname: Illies
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D048888:Fireflies / Q000201:enzymology; D007295:Inositol Phosphates / Q000378:metabolism*; D049409:Luciferases, Firefly / Q000378:metabolism; D049408:Luminescent Agents / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*
chemical_list: D007295:Inositol Phosphates; D049408:Luminescent Agents; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D049409:Luciferases, Firefly