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Photo-cross-linking and high-resolution mass spectrometry for assignment of RNA-binding sites in RNA-binding proteins

Open access
Kramer, Katharina, Sachsenberg, Timo, Beckmann, Benedikt M, Qamar, Saadia, Boon, Kum-Loong, Hentze, Matthias W, Kohlbacher, Oliver, Urlaub, Henning·2014·Nature Methods

Abstract

RNA-protein complexes play pivotal roles in many central biological processes. Although methods based on high-throughput sequencing have advanced our ability to identify the specific RNAs bound by a particular protein, there is a need for precise and systematic ways to identify RNA interaction sites on proteins. We have developed an experimental and computational workflow combining photo-induced cross-linking, high-resolution mass spectrometry and automated analysis of the resulting mass spectra for the identification of cross-linked peptides, cross-linking sites and the cross-linked RNA oligonucleotide moieties of such RNA-binding proteins. The workflow can be applied to any RNA-protein complex of interest or to whole proteomes. We applied the approach to human and yeast mRNA-protein complexes in vitro and in vivo, demonstrating its powerful utility by identifying 257 cross-linking sites on 124 distinct RNA-binding proteins. The open-source software pipeline developed for this purpose, RNPxl, is available as part of the OpenMS project.

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Metadata

Authors
Kramer, K., Sachsenberg, T., Beckmann, B. M., Qamar, S., Boon, K.-L., Hentze, M. W., Kohlbacher, O., & Urlaub, H.
Venue
Nature Methods
Published
2014-08-31
Volume
11
Issue
10
Pages
1064-1070
ISSN
  • 1548-7091 (ISSN-L)
  • 1548-7105
OpenAlex
W2077527436
Cited by
259
References
2
FWCI
10.34
Type
journal article
Language
en
Open access
bronze

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CoChat. (2026). Citation check: Photo-cross-linking and high-resolution mass spectrometry for assignment of RNA-binding sites in RNA-binding proteins. https://app.cochat.ai/papers/10.1038/nmeth.3092
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Kramer, K., Sachsenberg, T., Beckmann, B. M., Qamar, S., Boon, K.-L., Hentze, M. W., Kohlbacher, O., & Urlaub, H. (2014). Photo-cross-linking and high-resolution mass spectrometry for assignment of RNA-binding sites in RNA-binding proteins. Nature Methods. https://doi.org/10.1038/nmeth.3092

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