What is histone methyltransferase activity?
Histone methyltransferases (HMT) are histone-modifying enzymes (e.g., histone-lysine N-methyltransferases and histone-arginine N-methyltransferases), that catalyze the transfer of one, two, or three methyl groups to lysine and arginine residues of histone proteins.
What is histone methylation used for?
Histone methylation is important in modulating the accessibility of transcription factors to target genes and the subsequent changes in transcription. The site-specific methylation and demethylation of histone residues are catalyzed by methyltransferases and demethylases, respectively.
What is a lysine methyltransferase?
Lysine methyltransferases are enyzmes that catalyze the transfer of methyl groups from S-adenosylmethionine (SAM) to the lysine residues on histones, particularly histones H3 and H4. The dysregulation of this methylation is critical in the development of cancer.
What are the limitations of histone methyltransferase activity assays?
Histone methyltransferase activity assays Assay development for histone methyltransferases (HMTs) is difficult because they have often have low kcat (turnover is typically below 1 min -1) and low KM values for their methyl donor S-adenosylmethionine (SAM). HMT assays therefore have need to have very high sensitivity.
What is a histone modification assay?
In vitro histone modification (HM) assays are used to characterize the activity of chromatin-modifying enzymes. These assays provide information regarding the modification sites on histones, the product specificity, and the impact of other histone or nucleotide modifications on enzyme activity.
What do histone demethylase assays measure?
Histone demethylase assays usually measure the formation of formaldehyde, a by-product of demethylation. They are therefore susceptible to interference from detergents, thiol groups and a range of ions.
What are HMT activity assays and how are they used?
Many HMT activity assays measure S-adenosylhomocysteine (SAH) formation, a by-product of the methylation reaction. This means that they are not specific for any HMT, and can only be used with purified protein/protein complexes. They also often use 3 H-SAM as a donor, and so involve handling radioactive material.