Journal article
Simplified cysteine dioxygenase activity assay allows simultaneous quantitation of both substrate and product
E Siakkou, SM Wilbanks, GNL Jameson
Analytical Biochemistry | ACADEMIC PRESS INC ELSEVIER SCIENCE | Published : 2010
Abstract
A high-performance liquid chromatography (HPLC) method for enzyme activity assays using a hydrophilic interaction liquid chromatography (HILIC) column in combination with an evaporative light scattering detector was developed. The method was used to measure the activity of the non-heme mono-iron enzyme cysteine dioxygenase. The substrate cysteine and the product cysteine sulfinic acid are very weak chromophores, making direct ultraviolet (UV) detection without derivatization rather insensitive; moreover, derivatization of cysteine is often not efficient. Using the system described, underivatized substrate and product in samples from cysteine dioxygenase activity assays could be separated and..
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Awarded by Royal Society of New Zealand
Funding Acknowledgements
We thank Malcolm Rutledge (Department of Chemistry, University of Otago) for technical assistance during the purification of CDO protein. We also thank Mervyn Thomas (Department of Chemistry, University of Otago) for conducting the NMR experiments. This work was supported by the University of Otago Research Committee and the Royal Society of New Zealand Marsden Fund (09-UOO-069).