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 proteins from deconvolution and deisotoping through analysis 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 spectra and discuss the iterative use of the software to characterize proteoforms and to discover the sites of post-translational modifications.

Last modified

• 08/07/2017

Creator

• Paul M Thomas
• Caroline J DeHart
• Ryan T Fellers
• Luca Fornelli
• Neil L Kelleher

DOI

• https://doi.org/10.21985/N23T94

Publisher

• Springer

Alternate Identifier

• doi:10.21985/N2Q07P

Keyword

• ProSight Lite
• Top-down
• Bioinformatics
• Proteomics
• Mass spectrometry
• Intact protein

Date created

• 2017-02-02

Related url

•  https://link.springer.com/protocol/10.1007%2F978-1-4939-6783-4_18

Resource type

• Dataset

Source

• Protein Bioinformatics

License

• Attribution-NonCommercial-NoDerivs 3.0 United States