EZH2 (Y641N) TR-FRET Assay Kit

Catalog #: 52078
Size: 384 reactions
$1,450
Background:
EZH2 is the catalytic subunit of the PRC2/EED-EZH2 complex, which methylates Lys9 (H3K9me) and Lys27 (H3K27me) of histone H3, leading to transcriptional repression of the affected target gene. Able to mono-, di- and trimethylate Lys27 of histone H3 to form H3K27me1, H3K27me2 and H3K27me3, respectively. Compared to EZH2-containing complexes, it is more abundant in embryonic stem cells and plays a major role in forming H3K27me3, which is required for embryonic stem cell identity and proper differentiation. The PRC2/EED-EZH2 complex may also serve as a recruiting platform for DNA methyltransferases, thereby linking two epigenetic repression systems. Genes repressed by the PRC2/EED-EZH2 complex include HOXC8, HOXA9, MYT1, CDKN2A and retinoic acid target genes. EZH2 can also methylate non-histone proteins such as the transcription factor GATA4.
Description:
The EZH2 (Y641N) TR-FRET Assay Kit is designed to measure activity of the mutant EZH2 complex [EZH2 (Y641N)/EED/SUZ12/RbAp48/AEBP] in a homogeneous 384 reaction format. This FRET-based assay requires no time-consuming washing steps, making it especially suitable for high throughput screening applications. The key to the EZH2 (Y641N) Assay Kit is a highly specific antibody that recognizes methylated Histone H3K27. With this kit, only three simple steps on a microtiter plate are required for methyltransferase detection. First, Sadenosylmethionine is incubated with a sample containing assay buffer and methyltransferase enzyme for 4 hours. Next, antibody to methylated H3K27 is added. Finally, dye-labeled acceptor is added followed by fluorescence detection.
Synonym(s):
HTRF
Supplied As:
The EZH2 (Y641N) TR-FRET Assay Kit comes in a convenient format, with histone H3 peptide substrate, a Tb-labeled antibody against methylated K27 residue of Histone H3, S-adenosylmethionine, methyltransferase assay buffer, TR-FRET detection buffer, dye-lab
Format:

Instructions for use:
See datasheet for detailed protocol.
Storage / Stability:
At least 6 months from date of receipt when stored as directed.
Reference(s):

1. Dillon, S.C., et al. Genome Biology 2005; 6:227
2. Morin, R.D., et al. Nat Genet. 2010, 42(2):181

Notes:
MATERIALS OR INSTRUMENTS REQUIRED BUT NOT SUPPLIED:
Fluorescent microplate reader capable of measuring Time Resolved Fluorescence Resonance Energy Transfer (TR-FRET)
Adjustable micropipettor and sterile tips

This mutation is often cited in the scientific literature as Y641N in variant 3 (GenBank Accession No. NM_001203247).

Warning(s):
Avoid freeze/thaw cycles.
Scientific Category:
Methyltransferases
Product Type:
Assay Kits
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Product Datasheets

52078.pdf