Brief Communication

Identification of cross-linked peptides from complex samples

  • Nature Methods volume 9, pages 904906 (2012)
  • doi:10.1038/nmeth.2099
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Abstract

We have developed pLink, software for data analysis of cross-linked proteins coupled with mass-spectrometry analysis. pLink reliably estimates false discovery rate in cross-link identification and is compatible with multiple homo- or hetero-bifunctional cross-linkers. We validated the program with proteins of known structures, and we further tested it on protein complexes, crude immunoprecipitates and whole-cell lysates. We show that it is a robust tool for protein-structure and protein-protein–interaction studies.

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Acknowledgements

The authors wish to express gratitude to A.F. Hühmer and D. Horn (Thermo Fisher Scientific) for discussion; L.-L. Du (National Institute of Biological Sciences, Beijing (NIBS)) for critical suggestions; S.J. Lo (Chang Gung University) for the fib-1GFP strain; E.-Z. Shen, H.-Q. Wang, X.-D. He and G.-H. Cai (NIBS) for help with microscopy, GFP immunoprecipitation and peptide experiments; and R. Aebersold and T. Walzthöni (ETH Zurich) for help with xQuest search. We thank Z. Yuan, C. Liu, R.-X. Sun, Y. Fu and other members of the pFind team for discussion and support. This work was funded by the Ministry of Science and Technology of China (863 grant 2007AA02Z1A7 and 973 grant 2010CB835203 to M.-Q.D.; 863 grant 2008AA022310 and 973 grant 2010CB835402 to K.Y.; 973 grants 2012CB910602 and 2010CB912701 to S.-M.H.), the CAS Knowledge Innovation Program (KGCX1-YW-13) and an ICT basic research grant to S.-M.H. We thank the municipal government of Beijing for funds allocated to NIBS.

Author information

Author notes

    • Bing Yang
    • , Yan-Jie Wu
    • , Ming Zhu
    •  & Sheng-Bo Fan

    These authors contributed equally to this work.

Affiliations

  1. College of Biological Sciences, China Agricultural University, Beijing, China.

    • Bing Yang
  2. National Institute of Biological Sciences, Beijing, Beijing, China.

    • Bing Yang
    • , Ming Zhu
    • , Jinzhong Lin
    • , Shuang Li
    • , Yu-Xin Li
    • , Shu-Kun Luo
    • , Yue-He Ding
    • , She Chen
    • , Keqiong Ye
    •  & Meng-Qiu Dong
  3. Key Lab of Intelligent Information Processing, Institute of Computing Technology, Chinese Academy of Sciences, Beijing, China.

    • Yan-Jie Wu
    • , Sheng-Bo Fan
    • , Kun Zhang
    • , Hao Chi
    • , Hai-Feng Chen
    • , Le-Heng Wang
    • , Li-Yun Xiu
    •  & Si-Min He
  4. College of Life Sciences, Beijing Normal University, Beijing, China.

    • Ming Zhu
  5. Thermo Fisher Scientific, San Jose, California, USA.

    • Zhiqi Hao

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Contributions

B.Y. performed most wet-lab experiments and data analysis, and contributed to manuscript preparation; Y.-J.W. developed pLink; M.Z. performed peptide experiments (with B.Y.), non-BS3 cross-linking experiments and Y2H experiments; S.-B.F. fixed software bugs and maintained the system; J.L., S.L. and S.-K.L. purified the UTP-B, CNGP and F15E11.13-F15E11.14 protein complexes, respectively; K.Z. and L.-Y.X. developed pLabel for cross-links; H.C., H.-F.C. and L.-H.W. contributed to pLink development; Y.-X.L. contributed to data analysis; Y.-H.D. contributed to non-BS3 cross-linking experiments; Z.H. and S.C. contributed to MS analysis; K.Y. provided crucial samples and contributed to data interpretation and manuscript preparation; S.-M.H. directed software development and revised the manuscript; M.-Q.D. conceived the study, directed wet-lab experiments and wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Si-Min He or Meng-Qiu Dong.

Supplementary information

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    Supplementary Text and Figures

    Supplementary Figures 1–18, Supplementary Tables 1–20 and Supplementary Note