title: Substrate independent ATPase activity may complicate high throughput screening.
abstract: Inorganic phosphate release, [Pi], is often measured in an enzymatic reaction in a high throughput setting. Based on the published mechanism, we designed a protocol for our screening for inhibitors of SAICAR synthetase (PurC), and we found a gradual increase in [Pi] in positive control samples over the course of the day. Further investigation indicated that hydrolysis of ATP catalyzed by PurC, rather than substrate-related phosphate release, was responsible for a partial contribution to the signals in the control samples. Thus substrate-independent ATPase activity may complicate high throughput screening.
authors.affiliation: Department of Chemistry, University of Illinois at Chicago, Chicago, IL 60607, USA.
authors.forename: Micheal L
authors.initials: ML
authors.lastname: Tuntland
authors.affiliation: Department of Chemistry, University of Illinois at Chicago, Chicago, IL 60607, USA. Electronic address: lfung@uic.edu.
authors.forename: L W-M
authors.initials: LW
authors.lastname: Fung
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D004791:Enzyme Inhibitors / Q000737:chemistry*; D010453:Peptide Synthases / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D000251:Adenosine Triphosphatases; D010453:Peptide Synthases; C083513:PurC protein, Bacteria
keywords: High throughput screening; Malachite green; Substrate-independent ATPase activity