A follow-up to check the increased fluorescence signals at 0 min of the HDAC11 reaction with its substrate was performed. The data is uploaded on Zenodo.
Using Boc-Lys-(TFA)-AMC substrate, a preliminary time-course study was initiated to determine the appropriate HDAC11 concentration and the time to be used for the reaction (which are essential parameters to perform Km calculations later). Experimental details are on Zenodo.
In order to develop an assay for HDAC11, a kit-free method is being attempted wherein Boc-Lys-(TFA)-AMC, a previously reported substrate was used to check the activity of HDAC11 in a preliminary experiment. Dataset on Zenodo.
Preliminary tests to check if the Fluor-de-Lys-green HDAC activity kit could be used for developing a high-throughput HDAC11 assay have been performed here using controls and recombinantly purified HDAC11. Details on Zenodo.
HDAC 11 protein was purified from Sf9 cells using Ni-NTA chromatography. The protein was concentrated to 1.4 mg/ml (above which it tends to precipitate) and flash frozen. The details on the experimental procedures can be found on Zenodo.
Histone decaetylases (HDACs) are one of the several enzymes that take part in regulating gene expression. They catalyze the removal of acetyl groups from ε-N-acetylated lysines from both, histone and non-histone protein substrates. By doing this, they maintain a balance with respect to the activity of the acetyltransferases in the cells. The HDAC family consists Read More …