Efficient synthesis of 13C,15N-labeled RNA containing the cap structure m7GpppA

Hiroshi Matsuo, Tomohisa Moriguchi, Toshimitsu Takagi, Takahiro Kusakabe, Stephen Buratowski, Mitsuo Sekine, Yoshimasa Kyogoku, Gerhard Wagner

Research output: Contribution to journalArticlepeer-review

21 Citations (Scopus)

Abstract

Studying the mechanism by which the cap structure performs its numerous biological roles in mRNA metabolism by NMR spectroscopy requires the large- scale production of isotopically-labeled capped RNA. We present an efficient method for production of short RNA containing the cap structure m7GpppA, by combining chemical synthesis with enzymatic synthesis using a DNA primase. This method was employed to synthesize three capped RNA molecules, m7Gppp([13C,15N])A, m7Gppp([13C, 15N])ACC, and m7GpppA([13C,15N])C([13C,15N])C, that contain selective 13C,15N- labeled adenosine or cytosine nucleotides. This selective labeling technique enabled us to obtain the chemical shift assignments of these oligonucleotides in complex with the eukaryotic translation initiation factor 4E (eIF4E). Furthermore, we were able to observed intermolecular NOE interactions between the RNA and protein.

Original languageEnglish
Pages (from-to)2417-2421
Number of pages5
JournalJournal of the American Chemical Society
Volume122
Issue number11
DOIs
Publication statusPublished - Mar 22 2000

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

Fingerprint Dive into the research topics of 'Efficient synthesis of <sup>13</sup>C,<sup>15</sup>N-labeled RNA containing the cap structure m<sup>7</sup>GpppA'. Together they form a unique fingerprint.

Cite this