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A non-radioactive assay for selenophosphate synthetase activity using recombinant pyruvate pyrophosphate dikinase from Thermus thermophilus HB8.
Bioscience, Biotechnology, and Biochemistry 2016 October
Biosynthesis of selenocysteine-containing proteins requires monoselenophosphate, a selenium-donor intermediate generated by selenophosphate synthetase (Sephs). A non-radioactive assay was developed as an alternative to the standard [8-(14)C] AMP-quantifying assay. The product, AMP, was measured using a recombinant pyruvate pyrophosphate dikinase from Thermus thermophilus HB8. The KM and kcat for Sephs2-Sec60Cys were determined to be 26 μM and 0.352 min(-1), respectively.
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