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A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile
JoVE 杂志
免疫与感染
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JoVE 杂志 免疫与感染
A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile

A Purification and In Vitro Activity Assay for a (p)ppGpp Synthetase from Clostridium difficile

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09:53 min

November 03, 2018

DOI:

09:53 min
November 03, 2018

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Summary

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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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