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Optimization of mass spectrometry-compatible surfactants for shotgun proteomics

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Overview

authors

  • Chen, E. I.
  • Cociorva, D.
  • Norris, J. L.
  • Yates III, John

publication date

  • 2007

journal

  • Journal of Proteome Research  Journal

abstract

  • An optimization and comparison of trypsin digestion strategies for peptide/protein identifications by microLC-MS/MS with or without MS compatible detergents in mixed organic-aqueous and aqueous systems was carried out in this study. We determine that adding MS-compatible detergents to proteolytic digestion protocols dramatically increases peptide and protein identifications in complex protein mixtures by shotgun proteomics. Protein solubilization and proteolytic efficiency are increased by including MS-compatible detergents in trypsin digestion buffers. A modified trypsin digestion protocol incorporating the MS compatible detergents consistently identifies over 300 proteins from 5 microg of pancreatic cell lysates and generates a greater number of peptide identifications than trypsin digestion with urea when using LC-MS/MS. Furthermore, over 700 proteins were identified by merging protein identifications from trypsin digestion with three different MS-compatible detergents. We also observe that the use of mixed aqueous and organic solvent systems can influence protein identifications in combinations with different MS-compatible detergents. Peptide mixtures generated from different MS-compatible detergents and buffer combinations show a significant difference in hydrophobicity. Our results show that protein digestion schemes incorporating MS-compatible detergents generate quantitative as well as qualitative changes in observed peptide identifications, which lead to increased protein identifications overall and potentially increased identification of low-abundance proteins.

subject areas

  • Animals
  • Cell Extracts
  • Mass Spectrometry
  • Mice
  • Pancreas
  • Peptides
  • Proteomics
  • Solubility
  • Solutions
  • Solvents
  • Surface-Active Agents
  • Trypsin
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Identity

PubMed Central ID

  • PMC2570269

International Standard Serial Number (ISSN)

  • 1535-3893

Digital Object Identifier (DOI)

  • 10.1021/pr060682a

PubMed ID

  • 17530876
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Additional Document Info

start page

  • 2529

end page

  • 2538

volume

  • 6

issue

  • 7

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