Affiliations: Laboratory of Biochemistry, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
Note: [] To whom correspondence should be addressed: T.C. Stadtman, NHLBI/IR/LB, Building 3, Room 108, 3 Center Drive MSC 0320, Bethesda, MD 20892–0320, USA. Tel.: +1\,301\,496\,3002; Fax: +1\,301\,496\,0599.
Abstract: Selenophosphate synthetase catalyzes the formation of monoselenophosphate (SePO_{3}^{3-}) from ATP and selenide (reaction 1). \begin{equation} {\rm ATP + HSe^{-}} \rightarrow {\rm SePO_{3}^{3-} + PO_{4}^{3-} + AMP} \end{equation}
In one assay frequently used, [8‐^{14}C]AMP formation from [8‐^{14}C]ATP is estimated after separation of the nucleotides by thin‐layer 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‐^{32}P]ATP. The ^{32}P‐labeled selenophosphate is converted to [^{32}P]orthophosphate by treatment with iodine and estimated after removal of residual [^{32}P]ATP on charcoal.
Journal: Biofactors, vol. 6, no. 3, pp. 305-309, 1997
