1. A Universal Competitive Fluorescence Polarization Activity Assay for S-Adenosylmethionine Utilizing Methyltransferases. Tiffany L. Graves, Yi Zhang, and John E. Scott. Department of Pharmaceutical Sciences, Biomanufacturing Research Institute and Technology Enterprise, North Carolina Central University, Durham, NC 27707, USA. Anal Biochem. 2008 Feb15; 373(2):296-306. (Abstract).
SUMMARY: A competitive FPIA for SAH which utilizes an antibody to SAH and an SAH-fluorescein tracer. This assay has some limitations in detection level and assay window due to the fact that the antibody cross-reacts with SAM.
2. Development and Validation of a Generic Fluorescent Methyltransferase Activity Assay Based on the Transcreener AMP/GMP Assay. Klink, TA, Staeben M, Twesten, K, Kopp AL, Kumar M, Schall Dunn R, Pinchard C, Kleman KM, Klumpp M, Lowery RG, JBC 2011, submitted.
SUMMARY: Universal methyltransferase assay using coupling enzymes and fluorescent polarization immunoassay technology.
3. A Coupled Fluorescent Assay for Histone Methyltransferases. Collazo E, Couture JF, Bulfer S, Trievel RC., Department of Biological Chemistry, University of Michigan, Ann Arbor, MI 48109-0606, USA. Anal Biochem. 2005 Jul 1;342(1):86-92.(Abstract).
SUMMARY: A coupled enzyme assay for histone methyltransferase activity where SAH is converted to adenosine and homocysteine by SAH hydrolase, then the adenosine is removed from the reaction by adenosine deaminase (inosine is the product). The assay measures the resultant levels of sulfhydryl present from the homocysteine residues.
4. An Eenzyme-Coupled Continuous Spectrophotometric Assay for S-Adenosylmethionine-Dependent Methyltransferases.Dorgan KM, Wooderchak WL, Wynn DP, Karschner EL, Alfaro JF, Cui Y, Zhou ZS, Hevel JM. Department of Chemistry, Washington State University, Pullman, WA 99164, USA. Anal Biochem. 2006 Mar 15;350(2):249-55. (Abstract)
SUMMARY: This reference also describes a coupled enzymatic assay where SAH is converted to adenine (SAH nucleosidase), then adenine is deaminated to hypoxanthine which results in a drop in absorbance at 265nm. The detection of this drop in absorbance correlates with increased methyltransferase activity.
5. An Enzyme-Coupled Ultrasensitive Luminescence Assay for Protein Methyltransferases. Ibáñez G, McBean JL, Astudillo YM, Luo M. Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA. Anal Biochem. 2010 Jun 15;401(2):203-10. (Abstract).
SUMMARY: This paper describes a coupled three enzyme assay which converts SAH to adenine, AMP and finally ATP, which is detected and quantified with a luciferase-based detection module.
6. A Continuous Protein Methyltransferase (G9a) Assay for Enzyme Activity Measurement and Inhibitor Screening. Dhayalan A, Dimitrova E, Rathert P, Jeltsch A, J Biomol Screen 2009; 14:1129-1133.
SUMMARY: Radioactive Flashplate assay using native substrate with dose response curves using AdoHcy and BIX-01294.
7. Continuous Enzymatic Assay for Histone Lysine Methyltransferases. Rathert P, Cheng X, Jeltsch A, BioTechniques 2007; 43:602, 604, 606 passim.
SUMMARY: Streptavidin FlashPlate continuous peptide coated for Neurospora crassa Dim-5 histone H3 lysine 9 methyltransferase as a model system.
8. Small Molecule Regulators of Protein Arginine Methyltransferases. Cheng D, Yadav N, King RW, Swanson MS, Weinstein EJ, Bedford MT, J Biol Chem 2004; 279:23892-23899.
SUMMARY: Identification of small molecule inhibitors of histone methyltransferases including the selective arginine methyltransferase inhibitor AMI-1.
9. Reversal of H3K9me2 by a Small-Molecule Inhibitor for the G9a Histone Methyltransferase. Kubicek S, O’Sullivan RJ, August EM, Hickey ER, Zhang Q, Teodoro ML, Rea S, Mechtler K, Kowalski JA, Homon CA, Kelly TA, Jenuwein T, Mol Cell 2007; 25:473-481.
SUMMARY: Screening for G9a MT specific inhibitors using a 125,000 chemical library yielded seven hits, including identification of the biologically active inhibitor, BIX-01294 in both biochemical and cellular based assays.
10. Accessing Protein Methyltransferase and Demethylase Enzymology Using Microfluidic Capillary Electrophoresis. Wigle TJ, Provencher LM, Norris JL, Jin J, Brown PJ, Frye SV, Janzen WP, Chem Biol 2010 July 30; 17:695-704.
SUMMARY: Using UPLC-MS/MS (ultra-performance liquid chromatography-tandem MS), gel electrophoresis and thin-layer chromatography, the authors characterize the enzymatic activity of a protein arginine N-methyltransferase which selectively methylates histone H4.
11. Fluorescence-based High-Throughput Assay for Human DNA (Cytosine-5)-Methyltransferase 1. Ye Y, Stivers JT, Anal Biochem 2010 June 1; 401:168-172.
SUMMARY: Nonradioactive HTS assay method that is suitable for human DNA (cytosine-5)-methyltransferase 1 (DNMT1). The introduction of the methylation site into the recognition sequence of a restriction endonuclease, and the use of a fluorogenic read-out method are the key attributes of the technology.
A Fast and Efficient Method for Quantitative Measurement of S-Adenosyl-L-Methionine-Dependent Methyltransferase Activity With Protein Substrates.; Brenda B. Suh-Lailam and Joan M. Hevel. (Journal Abstract). Department of Chemistry and Biochemistry, Utah State University, Logan, UT 84322, USA and Center for Integrated Biosystems, Utah State University, Logan, UT 84322, USA. Anal Biochem. 2010 Mar 15; 398(2):218-24. (PubMed Abstract)
SUMMARY: Radioactive assay method for measuring protein methyltransferase activity where reverse-phase resin packed at the end of pipette tip is used to separate unreacted SAM from radiolabeled protein products.
Kinetic Analysis of Human Protein Arginine N-Methyltransferase 2: Formation of Monomethyl- and Asymmetric Dimethyl-Arginine Residues on Histone H4. Ted M. Lakowski and Adam Frankel., Division of Biomolecular and Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, BC, Canada, V6T 1Z3.Biochem. J. 2009 Jun 26; 421(2):253-61. (Full Text).
SUMMARY: Using UPLC-MS/MS (ultra-performance liquid chromatography-tandem MS), gel electrophoresis and thin-layer chromatography, the authors characterize the enzymatic activity of a protein arginine N-methyltransferase which selectively methylates histone H4.
A Nonradioactive DNA Methyltransferase Assay Adaptable to High-Throughput Screening. Youn-Hi Woo, P.T. Ravi Rajagopalan, and Stephen J. Benkovic. 414 Wartik Laboratory, Department of Chemistry, Pennsylvania State University, University Park, PA 16802, USA. Anal Biochem. 2005 Mar13; 340:336-340. (Abstract).
SUMMARY: An assay for DNA methlytransferases based upon protection from restriction endonucleases, then detection by an ELISA in microtiter plates.
An Enzyme-Coupled Colorimetric Assay for S-Adenosylmethionine-Dependent Methyltransferases.Hendricks CL, Ross JR, Pichersky E, Noel JP, Zhou ZS.Department of Chemistry, Carnegie Mellon University, Pittsburgh, PA 15213, USA. Anal Biochem. 2004 Mar 1;326(1):100-05. (Abstract)
SUMMARY: This paper describes a coupled enzyme assay which converts SAH to S-ribosylhomocysteine, then homocysteine. The thiol groups present in homocysteine are then detectable with a colorimetric assay.
A Real-Time Assay for CpG-Specific Cytosine-C5 Methyltransferase Activity. Wood RJ, McKelvie JC, Maynard-Smith MD, Roach PL. School of Chemistry, University of Southampton, Southampton, Hampshire, SO17 1BJ, UK. Nuc Acids Res. 2010 May; 38(9):e107. (Full Text)
SUMMARY: This paper describes an assay for monitoring DNA methyltransferase 1 (DNMT1) activity. An oligonucleotide is cleaved by a methylation sensitive endonuclease to free a fluorophore from its quencher and yield an increase in fluorescence.
Epigenetics: Tools and Technologies. Janzen WP, Wigle TJ, Jin J, Frye SV, Drug Discov Today Technol. 2010 Spring; 7(1):e59-e65; 7:e59-e65.
SUMMARY: Very informative and concise summary of the current state of epigenetic drug discovery with a focus on assay methods for methyltransferases. Highlights the regulation of chromatin, the complex of histone proteins, RNA and DNA that efficiently packages the genome, by specific modifications to histone proteins and DNA, and the recognition of these marks by other proteins and protein complexes.