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Straightforward ladder sequencing of peptides using a Lys-N metalloendopeptidase


We introduce a method for sequencing peptides by mass spectrometry using a metalloendopeptidase that cleaves proteins at the amino side of lysine (Lys-N). When analyzed by electron transfer dissociation (ETD)–based mass spectrometric sequencing, Lys-N–digested peptides that contain a single lysine residue produce spectra dominated by c-type fragment ions, providing simple ladders for sequence determination. This method should be a valuable strategy for de novo sequencing and the analysis of post-translational modifications.

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Figure 1: ETcaD peptide fragmentation spectra of doubly charged ions originating from Lys-C– and Lys-N–digested peptides.
Figure 2: Propensity of c-ion formation by ETcaD fragmentation for peptides originating from Lys-N and Lys-C digests.


  1. Aebersold, R. & Mann, M. Nature 422, 198–207 (2003).

    CAS  Article  Google Scholar 

  2. Witze, E.S., Old, W.M., Resing, K.A. & Ahn, N.G. Nat. Methods 4, 798–806 (2007).

    CAS  Article  Google Scholar 

  3. Standing, K.G. Curr. Opin. Struct. Biol. 13, 595–601 (2003).

    CAS  Article  Google Scholar 

  4. Zubarev, R.A., Kelleher, N.L. & McLafferty, F.W. J. Am. Chem. Soc. 120, 3265–3266 (1998).

    CAS  Article  Google Scholar 

  5. Syka, J.E.P., Coon, J.J., Schroeder, M.J., Shabanowitz, J. & Hunt, D.F. Proc. Natl. Acad. Sci. USA 101, 9528–9533 (2004).

    CAS  Article  Google Scholar 

  6. Molina, H., Horn, D.M., Tang, N., Mathivanan, S. & Pandey, A. Proc. Natl. Acad. Sci. USA 104, 2199–2204 (2007).

    CAS  Article  Google Scholar 

  7. Good, D.M., Wirtala, M., McAlister, G.C. & Coon, J.J. Mol. Cell. Proteomics 6, 1942–1951 (2007).

    CAS  Article  Google Scholar 

  8. Swaney, D.L. et al. Anal. Chem. 79, 477–485 (2007).

    CAS  Article  Google Scholar 

  9. Wysocki, V.H., Tsaprailis, G., Smith, L.L. & Breci, L.A. J. Mass Spectrom. 35, 1399–1406 (2000).

    CAS  Article  Google Scholar 

  10. Han, H., Xia, Y. & McLuckey, S.A. J. Proteome Res. 6, 3062–3069 (2007).

    CAS  Article  Google Scholar 

  11. Nonaka, T., Hashimoto, Y. & Takio, K. J. Biochem. 124, 157–162 (1998).

    CAS  Article  Google Scholar 

  12. Rao, K.C.S., Palamalai, V., Dunlevy, J.R. & Miyagi, M. Mol. Cell. Proteomics 4, 1550–1557 (2005).

    CAS  Article  Google Scholar 

  13. Peters, E.C., Horn, D.M., Tully, D.C. & Brock, A. Rapid Commun. Mass Spectrom. 15, 2387–2392 (2001).

    CAS  Article  Google Scholar 

  14. Beausoleil, S.A. et al. Proc. Natl. Acad. Sci. USA 101, 12130–12135 (2004).

    CAS  Article  Google Scholar 

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We thank A.F.M. Altelaar and B. van Breukelen for fruitful discussions and support. This work was supported by the Netherlands Proteomics Centre.

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Authors and Affiliations



N.T. and S.M. performed experiments; S.M. and A.J.R.H. designed experiments; N.T., A.J.R.H. and S.M. wrote the paper; M.M.D. analyzed peptide and fragment ion occurences.

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Correspondence to Albert J R Heck or Shabaz Mohammed.

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Supplementary Figures 1–4, Supplementary Tables 1–3, Supplementary Methods (PDF 451 kb)

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Taouatas, N., Drugan, M., Heck, A. et al. Straightforward ladder sequencing of peptides using a Lys-N metalloendopeptidase. Nat Methods 5, 405–407 (2008).

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