PatternLab: From Mass Spectra to Label‐Free Differential Shotgun Proteomics

Paulo C. Carvalho1, Juliana S. G. Fischer1, Tao Xu2, John R. Yates2, Valmir C. Barbosa3

1 Carlos Chagas Institute–Fiocruz, Paraná, Brazil, 2 Department of Cell Biology, The Scripps Research Institute, La Jolla, California, 3 Systems Engineering and Computer Science Program, COPPE, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil
Publication Name:  Current Protocols in Bioinformatics
Unit Number:  Unit 13.19
DOI:  10.1002/0471250953.bi1319s40
Online Posting Date:  December, 2012
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Abstract

PatternLab for proteomics is a self‐contained computational environment for analyzing shotgun proteomic data. Recent improvements incorporate modules to facilitate the computational analysis, such as FastaDBXtractor for sequence database preparation and ProLuCID runner for simplifying and managing the protein identification search engine; modules for pushing the limits on proteomics standards, such as SEPro, which relies on a semi‐labeled decoy approach for increasing confidence in filtering and organizing peptide spectrum matches; and modules with novel features, such as SEProQ for enabling label‐free quantitation by extracted ion chromatograms according to a distributed normalized ion abundance factor approach (dNIAF). Existing modules were also improved, such as the TFold module for pinpointing differentially expressed proteins. These new modules are integrated into the previously described arsenal of tools for further data analysis. Here we provide detailed instructions for operating and understanding them. Curr. Protoc. Bioinform. 40:13.19.1‐13.19.18. © 2012 by John Wiley & Sons, Inc.

Keywords: semi‐labeled decoy approach; filtering PSMs; dNIAF; quantitative proteomics; protein identification

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Preparing a Sequence Database to be Searched by ProLuCID or the Academic SEQUEST
  • Basic Protocol 2: Obtaining PSMs with ProLuCID and ProLuCID Runner
  • Basic Protocol 3: Filtering Results with the Search Engine Processor (SEPro)
  • Basic Protocol 4: Quantitating PSMs by dNIAFs with SEProQ
  • Basic Protocol 5: Using Regrouper to Port SEPro Spectral Counting or dNIAF Results to PatternLab
  • Basic Protocol 6: Using the Updated TFold Module for Pinpointing Differentially Expressed Proteins
  • Guidelines for Understanding Results
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

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Literature Cited

Literature Cited
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