Site-specific isotope labeling of long RNA for structural and mechanistic studies

Ikumi Kawahara, Kaichiro Haruta, Yuta Ashihara, Daichi Yamanaka, Mituhiro Kuriyama, Naoko Toki, Yoshinori Kondo, Kenta Teruya, Junya Ishikawa, Hiroyuki Furuta, Yoshiya Ikawa, Chojiro Kojima, Yoshiyuki Tanaka

研究成果: Contribution to journalArticle査読

8 被引用数 (Scopus)

抄録

A site-specific isotope labeling technique of long RNA molecules was established. This technique is comprised of two simple enzymatic reactions, namely a guanosine transfer reaction of group I self-splicing introns and a ligation with T4 DNA ligase. The trans-acting group I self-splicing intron with its external cofactor, 'isotopically labeled guanosine 5′-monophosphate' (5′-GMP), steadily gave a 5′-residue-labeled RNA fragment. This key reaction, in combination with a ligation of 5′-remainder non-labeled sequence, allowed us to prepare a site-specifically labeled RNA molecule in a high yield, and its production was confirmed with 15N NMR spectroscopy. Such a site-specifically labeled RNA molecule can be used to detect a molecular interaction and to probe chemical features of catalytically/structurally important residues with NMR spectroscopy and possibly Raman spectroscopy and mass spectrometry.

本文言語英語
ページ(範囲)e7
ジャーナルNucleic acids research
40
1
DOI
出版ステータス出版済み - 1 2012

All Science Journal Classification (ASJC) codes

  • Genetics

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