Articles
Development Of A Post-Reaction Derivatisation And Fluorescence Assay For 2-OG Dependent Dioxygenases
October 26, 2006
•Application Note: Development Of A Post-Reaction Derivatisation And Fluorescence Assay For 2-OG Dependent Dioxygenases
Members of the family of iron(II) and 2-oxoglutarate (2-OG) dependent dioxygenases fulfil a wide range of functions in natural systems. The most popular activity assay has previously relied on detecting the release of radiolabelled carbon dioxide from an appropriately labelled 2-OG molecule decarboxylated during the reaction. This procedure is time-consuming, tedious and is not easily converted for use in high-throughput or screening assays. The release of radioactive gas is also a point of concern.
This application note describes a novel assay that relies on postreaction derivatisation of residual 2-OG to determine the extent of enzyme reaction. Use of the NOVOstar both to perform the incubation and to detect the fluorescent product is explained.
The involvement of enzymes of the family in the hypoxic response system in animals has been characterised recently.2 Assay of both a prolyl hydroxylase enzyme and an asparaginyl hydroxylase enzyme is described.
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•Application Note: Development Of A Post-Reaction Derivatisation And Fluorescence Assay For 2-OG Dependent Dioxygenases
SOURCE: BMG LABTECH, GmbH
