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A Reader-Based Assay for mA Writers and Erasers


Wiedmer, Lars; Eberle, Stefanie Alexandra; Bedi, Rajiv Kumar; Śledź, Paweł; Caflisch, Amedeo (2019). A Reader-Based Assay for mA Writers and Erasers. Analytical Chemistry, 91(4):3078-3084.

Abstract

We have developed a homogeneous time-resolved fluorescence (HTRF)-based enzyme assay to measure the catalytic activity of N-methyladenosine (mA) methyltransferases and demethylases. The assay detects mA modifications using the natural mA-binding proteins (mA readers). The reaction product or substrate mA-containing RNA and the mA reader protein are fluorescently labeled such that their proximity during binding initiates Förster resonance energy transfer (FRET). We show that our HTRF assay can be used for high-throughput screening, which will facilitate the discovery of small-molecule modulators of mA (de)methylases.

Abstract

We have developed a homogeneous time-resolved fluorescence (HTRF)-based enzyme assay to measure the catalytic activity of N-methyladenosine (mA) methyltransferases and demethylases. The assay detects mA modifications using the natural mA-binding proteins (mA readers). The reaction product or substrate mA-containing RNA and the mA reader protein are fluorescently labeled such that their proximity during binding initiates Förster resonance energy transfer (FRET). We show that our HTRF assay can be used for high-throughput screening, which will facilitate the discovery of small-molecule modulators of mA (de)methylases.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:19 February 2019
Deposited On:06 Sep 2019 08:15
Last Modified:25 Sep 2019 00:44
Publisher:American Chemical Society (ACS)
ISSN:0003-2700
OA Status:Closed
Publisher DOI:https://doi.org/10.1021/acs.analchem.8b05500
PubMed ID:30715855

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