Chemical RNA digestion enables robust RNA-binding site mapping at single amino acid resolution

被引：34

作者：

Bae, Jong Woo ^{[1
,2
]}

Kwon, S. Chul ^{[1
,2
]}

Na, Yongwoo ^{[1
,2
]}

Kim, V. Narry ^{[1
,2
]}

Kim, Jong-Seo ^{[1
,2
]}

机构：

[1] Inst for Basic Sci Korea, Ctr RNA Res, Seoul, South Korea

[2] Seoul Natl Univ, Sch Biol Sci, Seoul, South Korea

来源：

NATURE STRUCTURAL & MOLECULAR BIOLOGY | 2020年 / 27卷 / 07期

关键词：

MASS-SPECTROMETRY; IDENTIFICATION; PHOSPHORYLATION; PROTEOME; PEPTIDE; DOMAINS;

D O I：

10.1038/s41594-020-0436-2

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

RNA-binding sites (RBSs) can be identified by liquid chromatography and tandem mass spectrometry analyses of the protein-RNA conjugates created by crosslinking, but RBS mapping remains highly challenging due to the complexity of the formed RNA adducts. Here, we introduce RBS-ID, a method that uses hydrofluoride to fully cleave RNA into mono-nucleosides, thereby minimizing the search space to drastically enhance coverage and to reach single amino acid resolution. Moreover, the simple mono-nucleoside adducts offer a confident and quantitative measure of direct RNA-protein interaction. Using RBS-ID, we profiled similar to 2,000 human RBSs and probedStreptococcus pyogenesCas9 to discover residues important for genome editing.

引用

页码：678 / +

页数：18

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