A Reader-Based Assay for m6A Writers and Erasers

被引：34
|
作者：
Wiedmer, Lars [1 ]
Eberle, Stefanie Alexandra [1 ]
Bedi, Rajiv Kumar [1 ]
Sledz, Pawel [1 ]
Caflisch, Amedeo [1 ]
机构：
[1] Univ Zurich, Dept Biochem, Winterthurerstr 190, CH-8057 Zurich, Switzerland
来源：
ANALYTICAL CHEMISTRY | 2019年 / 91卷 / 04期
基金：
瑞士国家科学基金会;
关键词：
STRUCTURAL BASIS; METTL3-METTL14; COMPLEX; NUCLEAR-RNA; METHYLATION; N6-METHYLADENOSINE; METTL3; METHYLTRANSFERASE; RECOGNITION; DEMETHYLASE; SUBUNIT;
D O I：
10.1021/acs.analchem.8b05500
中图分类号：
O65 [分析化学];
学科分类号：
070302 ; 081704 ;
摘要：
We have developed a homogeneous time resolved fluorescence (HTRF)-based enzyme assay to measure the catalytic activity of N-6-methyladenosine (m(6)A) methyltransferases and demethylases. The assay detects m(6)A modifications using the natural m(6)A-binding proteins (m(6)A readers). The reaction product or substrate m(6)A-containing RNA and the m(6)A reader protein are fluorescently labeled such that their proximity during binding initiates Forster resonance energy transfer (FRET). We show that our HTRF assay can be used for highthroughput screening, which will facilitate the discovery of smallmolecule modulators of m(6)A (de)methylases.
引用
收藏
页码：3078 / 3084
页数：7
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