Lessons in de novo peptide sequencing by tandem mass spectrometry

peer-reviewed · Mass Spectrometry Reviews · 2015

peer-reviewed · Mass Spectrometry Reviews · 2015. Katalin F. Medzihradszky et al. Mass spectrometry has become the method of choice for the qualitative and quantitative characterization of…
Date 2015-01-01
Type peer-reviewed
Venue Mass Spectrometry Reviews
Publisher Wiley
Contribution review
DOI 10.1002/mas.21406
Citations (OpenAlex) 211
Venue 2-year citedness 3.62

Abstract

Mass spectrometry has become the method of choice for the qualitative and quantitative characterization of protein mixtures isolated from all kinds of living organisms. The raw data in these studies are MS/MS spectra, usually of peptides produced by proteolytic digestion of a protein. These spectra are “translated” into peptide sequences, normally with the help of various search engines. Data acquisition and interpretation have both been automated, and most researchers look only at the summary of the identifications without ever viewing the underlying raw data used for assignments. Automated analysis of data is essential due to the volume produced. However, being familiar with the finer intricacies of peptide fragmentation processes, and experiencing the difficulties of manual data interpretation allow a researcher to be able to more critically evaluate key results, particularly because there are many known rules of peptide fragmentation that are not incorporated into search engine scoring. Since the most commonly used MS/MS activation method is collision-induced dissociation (CID), in this article we present a brief review of the history of peptide CID analysis. Next, we provide a detailed tutorial on how to determine peptide sequences from CID data. Although the focus of the tutorial is de novo sequencing, the lessons learned and resources supplied are useful for data interpretation in general.

Authors

  1. Katalin F. Medzihradszky · HUN-REN Biological Research Centre, University of California, San Francisco
  2. Robert J. Chalkley · University of California, San Francisco

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