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Straightforward ladder sequencing of peptides using a Lys-N metalloendopeptidase

Abstract

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.

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Acknowledgements

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

Authors

Contributions

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.

Corresponding authors

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). https://doi.org/10.1038/nmeth.1204

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