1. Structural Biology and Molecular Biophysics
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Trifunctional cross-linker for mapping protein-protein interaction networks and comparing protein conformational states

  1. Dan Tan
  2. Qiang Li
  3. Mei-Jun Zhang
  4. Chao Liu
  5. Chengying Ma
  6. Pan Zhang
  7. Yue-He Ding
  8. Sheng-Bo Fan
  9. Li Tao
  10. Bing Yang
  11. Xiangke Li
  12. Shoucai Ma
  13. Junjie Liu
  14. Boya Feng
  15. Xiaohui Liu
  16. Hong-Wei Wang
  17. Si-Min He
  18. Ning Gao
  19. Keqiong Ye
  20. Meng-Qiu Dong  Is a corresponding author
  21. Xiaoguang Lei
  1. Peking Union Medical College, Chinese Academy of Medical Sciences, China
  2. National Institute of Biological Sciences, China
  3. Institute of Computing Technology, Chinese Academy of Sciences, China
  4. Tsinghua University, China
  5. Chinese Academy of Medical Sciences, Peking Union Medical College, China
  6. Tianjin University, China
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Cite this article as: eLife 2016;5:e12509 doi: 10.7554/eLife.12509
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Abstract

To improve chemical cross-linking of proteins coupled with mass spectrometry (CXMS), we developed a lysine-targeted enrichable cross-linker containing a biotin tag for affinity purification, a chemical cleavage site to separate cross-linked peptides away from biotin after enrichment, and a spacer arm that can be labeled with stable isotopes for quantitation. By locating the flexible proteins on the surface of 70S ribosome, we show that this trifunctional cross-linker is effective at attaining structural information not easily attainable by crystallography and electron microscopy. From a crude Rrp46 immunoprecipitate, it helped identify two direct binding partners of Rrp46 and 15 protein-protein interactions (PPIs) among the co-immunoprecipitated exosome subunits. Applying it to E. coli and C. elegans lysates, we identified 3130 and 893 inter-linked lysine pairs, representing 677 and 121 PPIs. Using a quantitative CXMS workflow we demonstrate that it can reveal changes in the reactivity of lysine residues due to protein-nucleic acid interaction.

Article and author information

Author details

  1. Dan Tan

    Graduate Program, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  2. Qiang Li

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  3. Mei-Jun Zhang

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  4. Chao Liu

    Key Lab of Intelligent Information Processing of Chinese Academy of Sciences, Institute of Computing Technology, Chinese Academy of Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  5. Chengying Ma

    Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  6. Pan Zhang

    Graduate Program, Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  7. Yue-He Ding

    Graduate Program, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  8. Sheng-Bo Fan

    Key Lab of Intelligent Information Processing of Chinese Academy of Sciences, Institute of Computing Technology, Chinese Academy of Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  9. Li Tao

    Graduate Program, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  10. Bing Yang

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  11. Xiangke Li

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  12. Shoucai Ma

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  13. Junjie Liu

    Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  14. Boya Feng

    Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  15. Xiaohui Liu

    College of Pharmaceutical Science and Technology, Tianjin University, Tianjin, China
    Competing interests
    The authors declare that no competing interests exist.
  16. Hong-Wei Wang

    Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  17. Si-Min He

    Key Lab of Intelligent Information Processing of Chinese Academy of Sciences, Institute of Computing Technology, Chinese Academy of Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  18. Ning Gao

    Ministry of Education Key Laboratory of Protein Sciences, School of Life Sciences, Tsinghua University, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  19. Keqiong Ye

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.
  20. Meng-Qiu Dong

    Graduate Program, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China
    For correspondence
    dongmengqiu@nibs.ac.cn
    Competing interests
    The authors declare that no competing interests exist.
  21. Xiaoguang Lei

    National Institute of Biological Sciences, Beijing, China
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Brian Chait

Publication history

  1. Received: October 23, 2015
  2. Accepted: February 26, 2016
  3. Accepted Manuscript published: March 8, 2016 (version 1)
  4. Version of Record published: March 18, 2016 (version 2)
  5. Version of Record updated: September 20, 2016 (version 3)

Copyright

© 2016, Tan et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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