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The Eukaryotic RNA Exosome pp 383–400Cite as

Chemical Cross-Linking and Mass Spectrometric Analysis of the Endogenous Yeast Exosome Complexes

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 2062))

Abstract

Chemical cross-linking and mass spectrometric readout (CX-MS) has become a useful toolkit for structural analysis of protein complexes. CX-MS enables rapid detection of a larger number of cross-link peptides from the chemically cross-linked protein assembly, providing invaluable cross-link spatial restraints to understand the architecture of the complex. Since CX-MS is complementary with other structural and computational modeling tools, it can be used for integrative structural determination of large native protein assemblies. However, due to technical limitations, current CX-MS applications have still been predominantly confined to complexes reconstituted from recombinant proteins where large amount of purified materials are available. Cross-linking and hybrid structural proteomic analysis of endogenous protein complexes remains a challenge. In this chapter, we present a protocol that efficiently couples affinity capture of endogenous complexes with sensitive CX-MS analysis, with particular application to the yeast RNA processing exosome complexes.

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Acknowledgments

We thank Riccardo Pellarin (Institute of Pasteur) for preparing Fig. 4d; Junjie Wang (The Rockefeller University) for the development of CX-Circos.

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Authors and Affiliations

  1. Department of Cell Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

    Yufei Xiang, Zhuolun Shen & Yi Shi

  2. School of Medicine, Tsinghua University, Beijing, China

    Zhuolun Shen

Authors
  1. Yufei Xiang
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  2. Zhuolun Shen
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  3. Yi Shi
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Editors and Affiliations

  1. Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY, USA

    John LaCava

  2. Central European Institute of Technology (CEITEC), Masaryk University, Brno, Czech Republic

    Štěpánka Vaňáčová

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Xiang, Y., Shen, Z., Shi, Y. (2020). Chemical Cross-Linking and Mass Spectrometric Analysis of the Endogenous Yeast Exosome Complexes. In: LaCava, J., Vaňáčová, Š. (eds) The Eukaryotic RNA Exosome. Methods in Molecular Biology, vol 2062. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9822-7_18

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  • DOI: https://doi.org/10.1007/978-1-4939-9822-7_18

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9821-0

  • Online ISBN: 978-1-4939-9822-7

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