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Journal Article

Site-specific labeling of RNA at internal ribose hydroxyl groups: Terbium-assisted deoxyribozymes at work.

MPS-Authors
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Büttner,  L.
Research Group of Nucleic Acid Chemistry, MPI for Biophysical Chemistry, Max Planck Society;

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Javadi Zarnaghi,  F.
Research Group of Nucleic Acid Chemistry, MPI for Biophysical Chemistry, Max Planck Society;

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Höbartner,  C.
Research Group of Nucleic Acid Chemistry, MPI for Biophysical Chemistry, Max Planck Society;

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Fulltext (public)

2039333.pdf
(Publisher version), 3MB

Supplementary Material (public)

2039333_Suppl.pdf
(Supplementary material), 3MB

Citation

Büttner, L., Javadi Zarnaghi, F., & Höbartner, C. (2014). Site-specific labeling of RNA at internal ribose hydroxyl groups: Terbium-assisted deoxyribozymes at work. Journal of the American Chemical Society, 135(22), 8131-8137. doi:10.1021/ja503864v.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0019-DBC9-9
Abstract
A general and efficient single-step method was established for site-specific post-transcriptional labeling of RNA. Using Tb3+ as accelerating cofactor for deoxyribozymes, various labeled guanosines were site-specifically attached to 2'-OH groups of internal adenosines in in vitro transcribed RNA. The DNA-catalyzed 2',5'-phosphodiester bond formation proceeded efficiently with fluorescent, spin-labeled, biotinylated, or cross-linker-modified guanosine triphosphates. The sequence context of the labeling site was systematically analyzed by mutating the nucleotides flanking the targeted adenosine. Labeling of adenosines in a purine-rich environment showed the fastest reactions and highest yields. Overall, practically useful yields >70% were obtained for 13 out of 16 possible nucleotide (nt) combinations. Using this approach, we demonstrate preparative labeling under mild conditions for up to similar to 160-nt-long RNAs, including spliceosomal U6 small nuclear RNA and a cyclic-di-AMP binding riboswitch RNA.