A new (and freely available) original research article published ahead-of-print at SLAS Discovery Online describes a new methodology that enables the investigation of a large number of structurally conserved enzymes belonging to the Fe(II)/2-oxoglutarate-dependent dioxygenase superfamily. This superfamily comprises approximately 60 enzymes that are responsible for regulating a wide number of biological processes including epigenetic regulation, DNA/RNA repair and oxygen sensing. Of these, JumonjiC histone lysine demethylases (JMJCs) and prolyl hydroxylases are potential drug targets because of their relevance to human diseases.
The authors present an array of applications for their method. Using a miniaturized format, they characterize the enzymatic activity of several JumonjiC histone demethylases and other dioxygenases, for which high-throughput screening methods are currently unavailable. Using this simple succinate detection assay, the authors investigate substrate specificities and determine apparent kinetic parameters for different enzymes. They also validate the method as a screening tool for inhibitors using the LOPAC 1280 compound library and study the mode of action of selected hits against members of this enzyme family.
Bioluminescent succinate detection is resistant to chemical interference and it is effective for characterizing multiple Fe(II)/2-oxoglutarate-dependent dioxygenases with diverse substrate structures in a single format. By investigating a large number of these enzymes, this method could have a significant impact on the field of dioxygenase research.
