GCN5 Chemiluminescent Assay Kit
The GCN5 Chemiluminescent Assay Kit is an ELISA (enzyme-linked immunosorbent assay)-based assay designed to measure the histone acetyltransferase activity of GCN5 ((General control of amino acid synthesis, Yeast, Homolog)-like 2) for screening and profiling applications. The GCN5 assay kit comes in a convenient 96-well format, with enough recombinant purified GCN5 enzyme (amino acids 362-837), acetyl donor, pre-coated plate with histone substrate, all the reagents necessary for assay detection and blocking buffer for 96 enzyme reactions.
Figure 1: Schematic representation of the mechanism behind GCN5 Chemiluminescent Assay Kit.
GCN5 protein is added to a plate pre-coated with a histone substrate in the presence of acetyl-CoA. After washing, an anti-acetyl-lysine antibody is added, followed by a secondary HRP-antibody. Lastly, HRP substrate is added and the chemiluminescence signal generated can be measured. The signal is proportional to the acetyltransferase activity of GCN5
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- TBST buffer (1 x Tris-Buffered Saline, pH 8.0, containing 0.05% Tween-20)
- Luminometer or fluorescent microplate reader capable of reading chemiluminescence
- Adjustable micropipettor and sterile tips
- Rotating or rocker platform
| Catalog # | Name | Amount | Storage |
| 50074 | GCN5 (KAT2A), FLAG-Tag* | 5 µg | -80°C |
| 0.1 mM Acetyl-CoA | 10 µl | -20°C | |
| 79708 | 2x HAT Assay Buffer | 10 ml | -20°C |
| 52140U | Primary Antibody 21 | 12 µl | -20°C |
| 52131H | Secondary HRP-Labeled Antibody 2 | 10 µl | -20°C |
| 52100 | Blocking Buffer 4 | 50 ml | +4°C |
| 0.5M DTT | 200 µl | -20°C | |
| 79670 | ELISA ECL Substrate A (translucent bottle) | 6 ml | Room Temp |
| ELISA ECL Substrate B (brown bottle) | 6 ml | Room Temp | |
| White plate (or strips) pre-coated with Histone-Substrate | 1 plate | Room Temp |
*The concentration of the protein is lot-specific and will be indicated on the tube.
GCN5 ((General control of amino acid synthesis, Yeast, Homolog)-like 2), also known as KAT2A or lysine acetyltransferase 2A, can function as acetyltransferase, glutaryltransferase, succinyltransferase or malonyltransferase, depending on the cellular context, and it functions as a transcription regulator. As part of the SAGA (SPT-ADA-GCN5 acetyltransferase) and ATAC (ADA2A containing complex) complexes it functions as a histone acetyltransferase (HAT) on core histones. SAGA is a 19-subunit complex made of 4 units, including GCN5, that can recruit transcription proteins and modify promoter-proximal chromatin, being also involved in DNA damage repair and signaling pathways. ATAC can serve as a co-factor for c-Jun in the transcription of JNK (Jun N-terminal kinase) target genes. In the context of the hematopoietic lineage, GCN5 regulates maturation of NKT (natural killer T) cells, differentiation of granulocytic cells and proliferation of T cells. Both SAGA and ATAC have been linked to cancer and seem to be essential for cancer cell survival. A deep understanding of the GCN5 functions, and ability to inhibit or downregulate it may prove beneficial for patients suffering cancer where SAGA and ATAC are essential for cancer cell survival.
Herbst D., et al., 2021 Nature Structural & Molecular Biology 28: 989-996.
Suganuma T., et al., 2010 Cell 142 (5): 726-736.
Arede L., et al., 2022 Blood Adv. 6(1): 165-180.
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