An Optimized Microscale Thermophoresis Method for High-Throughput Screening of DNA Methyltransferase 2 Ligands
Robert A. Zimmermann, Marvin Schwickert, J. Laurenz Meidner, Zarina Nidoieva, Mark Helm, Tanja Schirmeister
Abstract
-adenosylhomocysteine derivative, 5-FAM-triazolyl-adenosyl-Dab, developed for a fluorescence polarization assay to investigate the histone methyltransferase mixed-lineage leukemia 1, we herein describe the applicability of this compound as a fluorescent tracer for the investigation of DNA-methyltransferase 2 (DNMT2), a human RNA methyltransferase. Based on these findings, we established a microscale thermophoresis (MST) assay for DNMT2. This displacement assay can circumvent various problems inherent to this method. Furthermore, we optimized a screening method via MST which even indicates if the detected binding is competitive and gives the opportunity to estimate the potency of a ligand, both of which are not possible with a direct binding assay.