Protein composition of human prespliceosomes isolated by a tobramycin affinity-selection method

Research output: Contribution to journalResearch articleContributedpeer-review

Contributors

  • Klaus Hartmuth - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute) (Author)
  • Henning Urlaub - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute) (Author)
  • Hans Peter Vornlocher - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute), Ribopharma AG (Author)
  • Cindy L. Will - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute) (Author)
  • Marc Gentzel - , European Molecular Biology Laboratory (EMBL) Heidelberg (Author)
  • Matthias Wilm - , European Molecular Biology Laboratory (EMBL) Heidelberg (Author)
  • Reinhard Lührmann - , Max Planck Institute for Biophysical Chemistry (Karl Friedrich Bonhoeffer Institute) (Author)

Abstract

Detailed knowledge of the composition and structure of the spliceosome and its assembly intermediates is a prerequisite for understanding the complex process of pre-mRNA splicing. To this end, we have developed a tobramycin affinity-selection method that is generally applicable for the purification of native RNP complexes. By using this method, we have isolated human prespliceosomes that are ideally suited for both biochemical and structural studies. MS identified >70 prespliceosome-associated proteins, including nearly all known U1 and U2 snRNP proteins, and expected non-snRNP splicing factors. In addition, the DEAD-box protein p68, RNA helicase A, and a number of proteins that appear to perform multiple functions in the cell, such as YB-1 and TLS, were detected. Several previously uncharacterized proteins of unknown function were also identified, suggesting that they play a role in splicing and potentially act during prespliceosome assembly. These data provide insight into the complexity of the splicing machinery at an early stage of its assembly.

Details

Original languageEnglish
Pages (from-to)16719-16724
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America : PNAS
Volume99
Issue number26
Publication statusPublished - 24 Dec 2002
Peer-reviewedYes
Externally publishedYes

External IDs

PubMed 12477934
ORCID /0000-0002-4482-6010/work/142251051

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