Methyltransferase Assay Services

Reaction Biology offers the largest collection of methyltransferase assays in the industry comprising over 30 assays for compound screening and profiling.

All methyltransferase assays are radioisotope based using 3H-AdoMet. MT HotSpotTM is a gold standard radioisotope based MT assay service available for both profiling and low-cost high-throughput screening. It is also the only methyltransferase assay technology that can use nucleosome, histone protein, and peptide as substrates without any modifications. All methyltransferase proteins and substrates are produced in house and are also available for purchase.

  • Free-choice screening and selectivity panel screening options available
  • Detection of inhibitors with varying binding modes
  • Low scale, large scale screening and high-throughput options as well as hit to lead screening options are available
  • Please ask us for assay development and research collaboration options

Protein Methyltransferase Enzyme Family

Protein methyltransferases (PMTs) are enzymes that aid in the transfer of methyl groups (—CH3) to specific sites on histones or acceptor proteins.

The phylogenetic tree shows the relationships between genes predicted to encode methyltransferases.

methyltransferases as targets for drug discovery, phylogenetic tree of methyltransferase family.

Assay Details

Assay Formats

MT HotSpotTM radiometric assay

methyltranferase assay principle for compound screening

Methyltransferases use tritium-labeled S-adenosyl-L-methionine (SAM) as the methyl donor that is converted to S-adenosyl-L-homocysteine (SAH) during the transfer of the radioactive methyl group to the histone substrate.

Reaction mixtures are incubated and spotted onto filter papers, which are then washed to remove unreacted SAM, leaving the bound radiolabeled product for detection.

Assay advantages

Due to the wash step requirement and the costs and safety concerns associated with the use of radioisotope materials, the radioisotope filter binding assay has traditionally been limited in its application to high-throughput screening. To address these limitations, Reaction Biology has developed the HotSpotTM assay technology, which is a biologically relevant, nanoliter-scale radioisotope filter binding platform, which offers high-throughput capacity and low material requirements.

The methyltransferase HotSpotTM assay requires no substrate modification, coupling enzymes, or detection antibodies.

Assay setup

Setups: Single-dose screening in duplicates or IC50 value determination with 5 or 10 concentrations. Other screening formats are available upon request.

Controls: No inhibitor (DMSO vehicle) control and for every assay, one target-specific control compound is tested in 10-dose IC50 format. 

Turnaround time: 10 business days for standard projects. Expedited scheduling and data delivery can be arranged prior to the commencement of the studies.

Panel screening: Screening on a panel of methyltransferases is performed on a regular basis every other week. Please see the next screening dates here.

Report: The raw data, % enzyme activity and control compound IC50 values will be reported in Excel format for single-dose assays. For IC50 orders, raw data, IC50 values, and curve fitting will be delivered in Excel format. Assay conditions, target, and substrate information are available upon request. Requirements for this information should be noted prior to the commencement of the study.

Screening facility: This assay is performed at our screening facility in Malvern, PA, USA.

Compound requirements: In brief, for a standard project, 20 µl of a 10 mM DMSO stock or solid material is needed. Less material is needed for large scale screening. Please refer to our FAQs for information regarding compound preparation and shipping.

High-Throughput Screening

Screen your own or our libraries for hit identification.

Available libraries:

  • Drug-like diversified small molecule libraries; > 110,000 compounds
  • Focused small libraries, including kinase inhibitors, FDA-approved inhibitors, epigenetic inhibitors, natural products, and a clinical collection.
  • Fragment libraries including a standard diversified fragment library, Fsp3-enriched fragments library, 3D fragment library, protein-protein interaction fragments, and covalent fragments library.

In addition to our high-throughput primary screening formats, we offer orthogonal screening formats and secondary screening formats. Hits can be characterized for their kinetic, binding affinity, mechanism of action, and intracellular activity.

Our medicinal chemistry partners are available to optimize the structure-activity-relationship of your hits to increase potency and reduce off-target and toxic effects and enhance the pharmacological profile of your lead compounds.

Assay Development

Methyltransferase assays that are not part of our portfolio can be developed in a custom-tailored fashion according to your research needs.

Our assay development service includes:

  • Plasmid construction, cDNA synthesis, and mutagenesis
  • E. coli or Baculovirus-based expression in insect cells
  • Co-expression with co-factors is optional
  • Purification via HIS, GST, GST-HIS, HALO-HIS, SUMO-HIS, TXN-HIS or other tags via affinity chromatography
  • Tag cleavage and enzyme activation are optional
  • Identification of suitable substrate
  • Determination of enzyme activity
  • Optimization of assay conditions