Epigenetics is one of the fastest growing fields in research because of its potential to identify therapeutic targets in oncology, immune and inflammatory diseases, cardiovascular disease, and personalized medicine. There are close to 500 epigenetic proteins that could be studied. These proteins are often referred to as writers, erasers, and readers.
The writers and erasers are the enzymes that add or remove the epigenetic modifications that lead to changes in gene expression. The writers are enzymes that methylate, acetylate, and phosphorylate substrates. The readers do as their name suggests: They read the modifications made by the writers. Examples of readers, also known as binding domains, are bromodomains, chromodomains, MBT domains, and tudor domains.
Until a few years ago, most epigenetic research was focused on histone deacetylases, more commonly known as HDACs. In the past five years, the focus has shifted quickly, and the evaluation of methyltransferase has became a high priority for scientists. These enzymes play a key role in modifying a large number of substrates such as histones and other proteins (e.g., p53), DNA, RNA, and small molecules making them key regulators of epigenetic states and, consequently, of gene expression.
The abundance of methyltransferases, their activity in effecting gene expression, and their potential in reversing modifications are key factors driving scientists to further understand their role and how they could be potential therapeutic targets.
The study of these important enzymes is proving to be challenging, however, because of their generally slow activity, environmental requirements such as buffers and pH balances, and variability in the substrates they modify as well as the number of modifications they make. All of these factors have made it difficult to develop universal assay approaches.
Methyltransferases are naturally slow acting enzymes and their reactions are known to take a significant period of time, making biologically relevant assays extremely difficult to develop. Several market surveys have reported that scientists are struggling to further their research and to understand these targets because manufacturers are not currently offering assays that meet their needs.
Until recently, scientists had been using a combination of various methods to study these enzymes such as ELISA, mass spec, Westerns, radioactivity, biomarker readouts, PCR, and several other technology platforms because there was not one platform besides radioactivity that scientists could use to screen all the methyltransferases.
The EPIgeneous Methyltransferase Assay from Cisbio was introduced to meet scientists’ needs and it is now being quickly adopted by scientists.
This mix-and-read assay uses HTRF technology to provide high sensitivity, flexibility, and is universal with regard to substrates, SAM concentrations, enzymatic buffers, and methyltransferases. This assay, which directly quantifies the methyltransferase reaction product S-adenosylhomocysteine (SAH), has been successfully validated on a variety of enzymes including G9a, SET8, NSD2, SET7/9, PRMT1, SETD2, DOT1L, and DNMT1. Its use has also been successfully demonstrated with all substrates (peptides, histones, nucleosomes, RNA, DNA, small molecules, and other proteins such as p53), making it a truly universal assay.