Brief Communication | Published:

Complementary positional proteomics for screening substrates of endo- and exoproteases

Nature Methods volume 7, pages 512515 (2010) | Download Citation

Abstract

We describe a positional proteomics approach to simultaneously analyze N- and C-terminal peptides and used it to screen for human protein substrates of granzyme B and carboxypeptidase A4 in human cell lysates. This approach allowed comprehensive proteome studies, and we report the identification of 965 database-annotated protein C termini, 334 neo–C termini resulting from granzyme B processing and 16 neo–C termini resulting from carboxypeptidase A4 processing.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    , , & Trends Cell Biol. 12, 146–150 (2002).

  2. 2.

    , , , & Anal. Biochem. 206, 344–352 (1992).

  3. 3.

    , , , & Nat. Methods 2, 193–200 (2005).

  4. 4.

    , , , & J. Proteome Res. 6, 4634–4645 (2007).

  5. 5.

    et al. Proteomics 8, 1362–1370 (2008).

  6. 6.

    , & Nat. Biotechnol. 19, 242–247 (2001).

  7. 7.

    et al. Mol. Cell. Proteomics 1, 376–386 (2002).

  8. 8.

    et al. Mol. Cell. Proteomics 8, 258–272 (2009).

  9. 9.

    , , , & Nat. Methods 6, 786–787 (2009).

  10. 10.

    et al. Cancer Res. 59, 2981–2988 (1999).

  11. 11.

    et al. Proc. Natl. Acad. Sci. USA 102, 3978–3983 (2005).

  12. 12.

    et al. Biochemistry 46, 6921–6930 (2007).

  13. 13.

    et al. Proteomics 5, 3537–3545 (2005).

  14. 14.

    et al. J. Biol. Chem. 279, 16907–16911 (2004).

  15. 15.

    et al. Nat. Methods 2, 771–777 (2005).

  16. 16.

    et al. Biochem. Biophys. Res. Commun. 261, 251–255 (1999).

  17. 17.

    et al. Mol. Cell. Proteomics 1, 896–903 (2002).

  18. 18.

    , , & J. Proteome Res. 6, 4770–4776 (2007).

  19. 19.

    et al. Mol. Cell. Proteomics 8, 2642–2652 (2009).

  20. 20.

    et al. BMC Proc. 3 (Suppl. 6), S6 (2009).

Download references

Acknowledgements

P. Bird (Monash University) kindly provided recombinant hGrB. P.V.D. was supported by a postdoctoral fellow grant of the Research Foundation, Flanders (FWO-Vlaanderen). K.H. is supported by a PhD grant of the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen). This work was supported by research grants from the Fund for Scientific Research, Flanders (Belgium) (project numbers G.0156.05, G.0077.06 and G.0042.07), the Concerted Research Actions (project BOF07/GOA/012) from Ghent University and the Interuniversity Attraction Poles (IUAP06). F.X.A. and S.B. acknowledge support from grants LSH6-2006-018830-CAMP (EC) and BIO2007-68046 (Ministerio de Ciencia y Innovación, Spain).

Author information

Affiliations

  1. Department of Medical Protein Research, Vlaams Instituut voor Biotechnologie, Ghent, Belgium.

    • Petra Van Damme
    • , An Staes
    • , Kenny Helsens
    • , Niklaas Colaert
    • , Evy Timmerman
    • , Joël Vandekerckhove
    •  & Kris Gevaert
  2. Department of Biochemistry, Ghent University, Ghent, Belgium.

    • Petra Van Damme
    • , An Staes
    • , Kenny Helsens
    • , Niklaas Colaert
    • , Evy Timmerman
    • , Joël Vandekerckhove
    •  & Kris Gevaert
  3. Institut de Biotecnologia i de Biomedicina and Departament de Bioquímica i Biologia Molecular, Universitat Autònoma de Barcelona, Bellaterra, Spain.

    • Silvia Bronsoms
    •  & Francesc X Aviles

Authors

  1. Search for Petra Van Damme in:

  2. Search for An Staes in:

  3. Search for Silvia Bronsoms in:

  4. Search for Kenny Helsens in:

  5. Search for Niklaas Colaert in:

  6. Search for Evy Timmerman in:

  7. Search for Francesc X Aviles in:

  8. Search for Joël Vandekerckhove in:

  9. Search for Kris Gevaert in:

Contributions

P.V.D., S.B., F.X.A., J.V. and K.G. designed the experiments. P.V.D., A.S. and E.T. generated the data. P.V.D., K.H. and N.C. analyzed the data. P.V.D., F.X.A., J.V. and K.G. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Petra Van Damme or Kris Gevaert.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–3 and Supplementary Tables 2–3

Excel files

  1. 1.

    Supplementary Table 1

    General overview of peptide and protein identifications.

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nmeth.1469

Further reading