Continuous enzymatic assay for histone lysine methyltransferases

Abstract

We describe a continuous peptide methylation assay using the Neurospora crassa Dim-5 histone H3 lysine 9 (H3K9) methyltransferase as a model system. The assay uses streptavidin FlashPlates coated with target peptide. Since no washing and pipeting steps were required after the addition of the enzyme/S-adenosyl-l-methionine (AdoMet) mixture to the microplate, a continuous readout of the reaction progress was possible. We show that this assay is highly reproducible (with errors in the order of ±3%). The continuous assay is well suited for the simultaneous analysis of up to 384 samples, thus allowing for a rapid screening of methylation rates of different substrates under different conditions or in the presence of inhibitors.