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Applications: Study enzyme kinetics of p300; Screen small molecular inhibitors for drug discovery and High throughput (HTS) applications.
Background: Histone acetyltransferase p300, also known as p300 HAT or EP300, is a histone acetyltransferase that regulates gene transcription through chromatin remodeling. Therefore, p300 plays an important role in the regulation of cell proliferation or differentiation. p300 is closely related to CREB binding protein. It interacts with many transcription factors. It also mediates cAMP-gene regulation by binding specifically to the phosphorylated CREB protein. Mutations in p300 are involved in the rare genetic disease Rubinstein–Taybi syndrome and in several types of cancer, including stomach, colon, breast, and pancreatic cancer.
Contraindications: DMSO concentrations above 0.5%.Green and blue dyes that absorb light in the AlphaScreen® signal emission range (520-620 nm), such as Trypan Blue.Avoid the use of the potent singlet oxygen quenchers such as sodium azide (NaN3) or metal ions (Fe2+, Fe3+, Cu2+, Zn2+ and Ni2+).The presence of >1% RPMI 1640 culture medium leads to a signal reduction due to the presence of excess biotin and iron in this medium. MEM, which lacks these components, does not affect AlphaScreen®
Description: The p300 Homogeneous Assay Kit contains an AlphaLISA® assay designed for screening inhibitors of p300 in a convenient 384-well format. This efficient method requires no time-consuming washing steps. Only a few steps are needed. The assay is based on the enzymatic transfer of an acetyl group from acetyl CoA to a specific lysine residue within a biotinylated peptide substrate. After incubation with p300, the sample is incubated with a combination of an antibody that specifically binds the acetylated peptide and acceptor beads that bind the antibody. This is followed by incubation with streptavidin-labeled donor beads and reading of the Alpha-counts.
Storage Stability: This assay kit will perform optimally for up to 12 months from date of receipt when the materials are stored as directed.
Supplied As: The p300 Homogeneous Assay Kit comes in a convenient AlphaLISA® format, with biotinylated histone substrate, Acetyl CoA, primary antibody, assay buffers, and purified p300 for 384 enzyme reactions.
Uniprot: Q09472
Warnings: Avoid freeze/thaw cycles.
Biosafety Level: Not applicable (BSL-1)
References: Trievel, R. C., et al. (2000). Anal. Biochem. 287(2):319-28.
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