化学
核糖核酸
N6-甲基腺苷
突变
酶
脱甲基酶
假尿苷
生物化学
核苷
寡核苷酸
计算生物学
立体化学
生物物理学
DNA
基因
甲基转移酶
转移RNA
突变
生物
表观遗传学
甲基化
作者
Florian Seitz,Tina Jungnickel,Nicole Kleiber,Jens Kretschmer,Julia Dietzsch,Juliane Adelmann,Katherine E. Bohnsack,Markus T. Bohnsack,Claudia Höbartner
摘要
N6-methyladenosine (m6A) is an important modified nucleoside in cellular RNA associated with multiple cellular processes and is implicated in diseases. The enzymes associated with the dynamic installation and removal of m6A are heavily investigated targets for drug research, which requires detailed knowledge of the recognition modes of m6A by proteins. Here, we use atomic mutagenesis of m6A to systematically investigate the mechanisms of the two human m6A demethylase enzymes FTO and ALKBH5 and the binding modes of YTH reader proteins YTHDF2/DC1/DC2. Atomic mutagenesis refers to atom-specific changes that are introduced by chemical synthesis, such as the replacement of nitrogen by carbon atoms. Synthetic RNA oligonucleotides containing site-specifically incorporated 1-deaza-, 3-deaza-, and 7-deaza-m6A nucleosides were prepared by solid-phase synthesis and their RNA binding and demethylation by recombinant proteins were evaluated. We found distinct differences in substrate recognition and transformation and revealed structural preferences for the enzymatic activity. The deaza m6A analogues introduced in this work will be useful probes for other proteins in m6A research.
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