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A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile
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A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile
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November 03, 2018
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Here, we describe a method for purifying histidine-tagged pyrophosphokinase enzymes and utilizing thin layer chromatography of radiolabelled substrates and products to assay for the enzymatic activity in vitro. The enzyme activity assay is broadly applicable to any kinase, nucleotide cyclase, or phosphor-transfer reaction whose mechanism includes nucleotide triphosphate hydrolysis.
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Pokhrel, A., Poudel, A., Purcell, E. B. A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile. J. Vis. Exp. (141), e58547, doi:10.3791/58547 (2018).
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