KDM6A/KDM6B Activity Quantification Assay Kit (Fluorometric)

KDM6A/KDM6B Activity Quantification Assay Kit (Fluorometric) (ab156911) is a complete set of optimized reagents, designed for measuring activity/inhibition of KDM6A and KDM6B using nuclear extracts or purified enzymes from a broad range of species such as mammals, plants, fungi, and bacteria, in a variety of forms including, but not limited to cultured cells and fresh and frozen tissues. There are currently a limited number of methods used for detecting KDM6A/KDM6B activity/inhibition.

The traditional method is based on the measurement of formaldehyde release, a by-product of KDM6A/KDM6B enzymatic reaction, and has significant weaknesses: (1) large amounts (at μg level) of substrate and enzyme are required; (2) nuclear extracts from cell/tissues cannot be used; (3) redox-sensitive KDM6A/KDM6B inhibitiors are not suitable for testing with such methods; (4) high intereference by SDS, DMSO, thiol-containing chemicals, and ions, which are often contained in enzyme solutions, tested compound solvents, and assay buffers; and (5) less accurate than direct measurement of KDM6A/KDM6B-converted demethylated products.

Lysine specific demethylase 6A (KDM6A/UTX) and lysine specific demethylase 6B (KDM6B/JMJD3) demethylase lysine 27 on histone H3, thereby playing a central role in histone code. KDM6A and KDM6B are JmjC (Jumonji)-domain-containing proteins and catalyze the removal of di- and tri-methylation from histone H3-K27 by using a hydroxylation reaction with iron and alpha-glutarate as cofactors.