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Bioinformatics Analysis of Top-Down Mass Spectrometry Data with ProSight Lite

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Protein Bioinformatics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1558))

Abstract

Traditional bottom-up mass spectrometry-based proteomics relies on the use of an enzyme, often trypsin, to generate small peptides (typically < 25 amino acids long). In top-down proteomics, proteins remain intact and are directly measured within the mass spectrometer. This technique, while inherently simpler than bottom-up proteomics, generates data which must be processed and analyzed using software tools “purpose-built” for the job. In this chapter, we will show the analysis of intact protein spectra through deconvolution, deisotoping, and searching with ProSight Lite, a free, vendor-agnostic tool for the analysis of top-down mass spectrometry data. We will illustrate with two examples of intact protein fragmentation spectra and discuss the iterative use of the software to characterize proteoforms and discover the sites of post-translational modifications.

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Acknowledgments

The authors are grateful to members of the Kelleher Research Group/Proteomics Center of Excellence for helpful discussions, particularly Joseph Greer, Richard LeDuc, and Bryan Early. This work was supported by Award No. P41GM108569 from the National Institute of General Medical Sciences The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

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Authors and Affiliations

  1. Departments of Chemistry, Molecular Biosciences, the Proteomics Center of Excellence and the Robert H. Lurie Comprehensive Cancer Center at the Feinberg School of Medicine, Northwestern University, Evanston, IL, 60208, USA

    Caroline J. DeHart, Ryan T. Fellers, Luca Fornelli, Neil L. Kelleher & Paul M. Thomas

Authors
  1. Caroline J. DeHart
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  2. Ryan T. Fellers
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  3. Luca Fornelli
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  4. Neil L. Kelleher
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  5. Paul M. Thomas
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Corresponding author

Correspondence to Paul M. Thomas .

Editor information

Editors and Affiliations

  1. Center for Bioinformatics and Computational Biology, University of Delaware, Newark, Delaware, USA

    Cathy H. Wu

  2. Center for Bioinformatics and Computational Biology, University of Delaware, Newark, Delaware, USA

    Cecilia N. Arighi

  3. Department of Biochemistry and Molecular and Cellular Biology, Georgetown University Medical Center, Washington, District of Columbia, USA

    Karen E. Ross

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DeHart, C.J., Fellers, R.T., Fornelli, L., Kelleher, N.L., Thomas, P.M. (2017). Bioinformatics Analysis of Top-Down Mass Spectrometry Data with ProSight Lite. In: Wu, C., Arighi, C., Ross, K. (eds) Protein Bioinformatics. Methods in Molecular Biology, vol 1558. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6783-4_18

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  • DOI: https://doi.org/10.1007/978-1-4939-6783-4_18

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6781-0

  • Online ISBN: 978-1-4939-6783-4

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