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A non-radioactive and two radioactive assays for selenophosphate synthetase activity
1Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Abstract:
Selenophosphate synthetase catalyzes the formation of monoselenophosphate (SePO3(3-)) from ATP and selenide (reaction 1). [formula: see text] In one assay frequently used, [8-14C]AMP formation from [8-14C]ATP is estimated after separation of the nucleotides by thinlayer chromatography. An alternative non-radioactive assay in which the AMP product is estimated using AMP deaminase is described. The highly oxygen-labile selenophosphate product can be estimated in an assay employing [gamma-32P]ATP. The 32P-labeled selenophosphate is converted to [32P]orthophosphate by treatment with iodine and estimated after removal of residual [32P]ATP on charcoal.