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Expressed In: Baculovirus Protein Species: Human
Contents
A representative Technical Data Sheet (TDS) is provided here. Please refer to the lot-specific TDS you will receive with your order for the lot-specific buffer contents and protein concentration.
Background
KDM6A (lysine (K)-specific demethylase 6A), also known as UTX (Ubiquitously Transcribed Tetratricopeptide Repeat, X Chromosome) is a protein that functions as a histone demethylase that preferentially demethylates di- and trimethylated lysine 27 residues of histone H3, while it has no activity on monomethylated H3K27 residues. KDM6A plays a central role in posterior development by regulating HOX gene expression. Demethylation of lysine 27 is concomitant with methylation of lysine 4 of histone H3 and regulates the recruitment of the PRC1 complex and monoubiquitination of histone H2A.
Application Notes
Recombinant UTX / KDM6A is suitable for use in the study of enzyme kinetics, inhibitor screening, and selectivity profiling.
Histone Demethylase Assay Conditions:
50 mM HEPES pH 7.5, 0.02% Triton X100, 100 μM 2OG, 100 μM Ascorbate, 50 μM (NH4)2Fe(SO4)2·6H2O, 1 mM TCEP, 100 nM Recombinant UTX / KDM6A protein, and 3.3 μM H3K27me3 peptide at 2 hours at room temperature. MALDI-TOF was used for detection.
Protein Details
Recombinant UTX / KDM6A (accession number NP_001278345.1) was expressed in Sf9 cells and contains an N-terminal FLAG-Tag with an observed molecular weight of 158.8 kDa. The recombinant protein is >72% pure by SDS-PAGE.
Recombinant UTX / KDM6A protein gel.
UTX / KDM6A protein was run on an SDS-PAGE gel and stained with Coomassie blue.
UTX / KDM6A activity assay.
Recombinant UTX / KDM6A activity measured using a demethylation assay. MALDI-TOF was used for detection.
UTX / KDM6A AlphaLISA assay.
UTX / KDM6A enzyme was used in an AlphaLISA assay to determine enzyme linearity, 2OG Km and IC50 of two reference compounds. 10 nM-20 nM KDM6A was used to develop an HTS screening assay and well-to-well reproducibility was also assessed at Z’=0.64. Demethylated peptide (H3K27me2) was measured using methylation-specific antibody. This data was generated and kindly provided courtesy of ChemPartner.
Storage
Recombinant proteins in solution are temperature sensitive and must be stored at -80°C to prevent degradation. Avoid repeated freeze/thaw cycles and keep on ice when not in storage.
Guarantee
This product is guaranteed for 6 months from date of receipt.
This product is for research use only and is not for use in diagnostic procedures.
