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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Singh, P., Panchaud, A. & Goodlett, D.R. Anal. Chem. 82, 2636–2642 (2010).
Zhang, H. et al. Mol. Cell. Proteomics 8, 409–420 (2009).
Petrotchenko, E.V., Serpa, J.J. & Borchers, C.H. Mol. Cell. Proteomics 10, M110.001420 (2011).
Kao, A. et al. Mol. Cell. Proteomics 10, M110.002212 (2011).
Hermanson, G.T. Bioconjugate Techniques 2nd edn Ch. 4, 241–243 (Academic, London, 2008).
Li, S. et al. Genes Dev. 25, 2409–2421 (2011).
Dragon, F. et al. Nature 417, 967–970 (2002).
Grandi, P. et al. Mol. Cell 10, 105–115 (2002).
Krogan, N.J. et al. Mol. Cell 13, 225–239 (2004).
Champion, E.A., Lane, B.H., Jackrel, M.E., Regan, L. & Baserga, S.J. Mol. Cell. Biol. 28, 6547–6556 (2008).
Lee, L.W., Lo, H.W. & Lo, S.J. Gene 455, 16–21 (2010).
Aittaleb, M. et al. Nat. Struct. Biol. 10, 256–263 (2003).
Lin, J. et al. Nature 469, 559–563 (2011).
Rinner, O. et al. Nat. Methods 5, 315–318 (2008).
Xu, H., Hsu, P.H., Zhang, L., Tsai, M.D. & Freitas, M.A. J. Proteome Res. 9, 3384–3393 (2010).
Su, C. et al. Nucleic Acids Res. 36, D632–D636 (2008).
Kubala, M.H., Kovtun, O., Alexandrov, K. & Collins, B.M. Protein Sci. 19, 2389–2401 (2010).
Panchaud, A., Singh, P., Shaffer, S.A. & Goodlett, D.R. J. Proteome Res. 9, 2508–2515 (2010).
Brenner, S. Genetics 77, 71–94 (1974).
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-1∷GFP 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.
The authors declare no competing financial interests.
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Yang, B., Wu, YJ., Zhu, M. et al. Identification of cross-linked peptides from complex samples. Nat Methods 9, 904–906 (2012). https://doi.org/10.1038/nmeth.2099
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