Protocol | Published:

Analyzing trapped protein complexes by Virotrap and SFINX

Nature Protocols volume 12, pages 881898 (2017) | Download Citation

Abstract

The analysis of protein interaction networks is one of the key challenges in the study of biology. It connects genotypes to phenotypes, and disruption of such networks is associated with many pathologies. Virtually all the approaches to the study of protein complexes require cell lysis, a dramatic step that obliterates cellular integrity and profoundly affects protein interactions. This protocol starts with Virotrap, a novel approach that avoids the need for cell homogenization by fusing the protein of interest to the HIV-1 Gag protein, trapping protein complexes in virus-like particles. By using the straightforward filtering index (SFINX), which is a powerful and intuitive online tool (http://sfinx.ugent.be) that enables contaminant removal from candidate lists resulting from mass-spectrometry-based analysis, we provide a complete workflow for researchers interested in mammalian protein complexes. Given direct access to mass spectrometers, researchers can process up to 24 samples in 7 d.

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Acknowledgements

K.T. is a PhD student with the Agency for Innovation by Science and Technology (IWT). K.G. acknowledges support from the Fund for Scientific Research-Flanders (FWO-Vlaanderen, grant G011312N) and the Ghent University Concerted Research Actions (grant BOF14/GOA/013). J.T. was supported by grants from IUAP P6/36, the GROUP-ID MRP-UGent and the Fund for Scientific Research-Flanders (grants G.0747.10N and G.0864.10), and is a recipient of a European Research Council (ERC) Advanced Grant (Cytokine Receptor Signaling Revisited, 340941). S.E. was supported by a Methusalem grant to J.T.

Author information

Author notes

    • Kevin Titeca
    •  & Emmy Van Quickelberghe

    These authors contributed equally to this work.

Affiliations

  1. VIB-UGent Center for Medical Biotechnology, Ghent, Belgium.

    • Kevin Titeca
    • , Emmy Van Quickelberghe
    • , Noortje Samyn
    • , Delphine De Sutter
    • , Annick Verhee
    • , Kris Gevaert
    • , Jan Tavernier
    •  & Sven Eyckerman
  2. Department of Biochemistry, Ghent University, Ghent, Belgium.

    • Kevin Titeca
    • , Emmy Van Quickelberghe
    • , Noortje Samyn
    • , Delphine De Sutter
    • , Annick Verhee
    • , Kris Gevaert
    • , Jan Tavernier
    •  & Sven Eyckerman

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Contributions

The Virotrap protocol was developed by S.E., E.V.Q. and K.T. with excellent assistance from N.S., A.V. and D.D.S., and support and advice from K.G. and J.T. SFINX was developed by K.T. under the guidance of J.T. and S.E. The manuscript was written by K.T., E.V.Q. and S.E., with corrections from K.G. and J.T.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Sven Eyckerman.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Figures and Tables

    Supplementary Figures 1–7.

CSV files

  1. 1.

    Supplementary Data 1

    Example file for 'Basic data' input (semicolon-separated).

Text files

  1. 1.

    Supplementary Data 2

    Example file for 'Bait identities' input.

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DOI

https://doi.org/10.1038/nprot.2017.014

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