Optimization of High-Throughput Methyltransferase
Assays for the Discovery of Small Molecule Inhibitors
Posted on 2020-06-27 - 16:03
Methyltransferases
(MTases) play diverse roles in cellular processes. Aberrant methylation
levels have been implicated in many diseases, indicating the need
for the identification and development of small molecule inhibitors
for each MTase. Specific inhibitors can serve as probes to investigate
the function and validate therapeutic potential for the respective
MTase. High-throughput screening (HTS) is a powerful method to identify
initial hits for further optimization. Here, we report the development
of a fluorescence-based MTase assay and compare this format with the
recently developed MTase-Glo luminescence assay for application in
HTS. Using protein N-terminal methyltransferase 1 (NTMT1) as a model
system, we miniaturized to 1536-well quantitative HTS format. Through
a pilot screen of 1428 pharmacologically active compounds and subsequent
validation, we discovered that MTase-Glo produced lower false positive
rates than the fluorescence-based MTase assay. Nevertheless, both
assays displayed robust performance along with low reagent requirements
and can potentially be employed as general HTS formats for the discovery
of inhibitors for any MTase.
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Dong, Guangping; Yasgar, Adam; Peterson, Darrell L.; Zakharov, Alexey; Talley, Daniel; Cheng, Ken Chih-Chien; et al. (2020). Optimization of High-Throughput Methyltransferase
Assays for the Discovery of Small Molecule Inhibitors. ACS Publications. Collection. https://doi.org/10.1021/acscombsci.0c00077
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AUTHORS (9)
GD
Guangping Dong
AY
Adam Yasgar
DP
Darrell L. Peterson
AZ
Alexey Zakharov
DT
Daniel Talley
KC
Ken Chih-Chien Cheng
AJ
Ajit Jadhav
AS
Anton Simeonov
RH
Rong Huang
KEYWORDS
Small Molecule Inhibitors Methyltra...High-Throughput Methyltransferase A...High-throughput screeningAberrant methylation levelsreagent requirements1428 pharmacologicallyHTS formatfluorescence-based MTase assayHTS formatsMTase-Glo luminescence assaypilot screenmolecule inhibitorsmodel systemNTMTprotein N-terminal methyltransferase 1Specific inhibitors