Evaluation of different multidimensional LC-MS/MS pipelines for isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomic analysis of potato tubers in response to cold storage

J Proteome Res. 2011 Oct 7;10(10):4647-60. doi: 10.1021/pr200455s. Epub 2011 Sep 6.

Abstract

Cold-induced sweetening in potato tubers is a costly problem for the food industry. To systematically identify the proteins associated with this process, we employed a comparative proteomics approach using isobaric, stable isotope coded labels to compare the proteomes of potato tubers after 0 and 5 months of storage at 5 °C. We evaluated both high pH reverse phase (hpRP) liquid chromatography (LC) and off-gel electrophoresis (OGE) as first dimension fractionation methods followed by nanoLC-MS/MS, using two high performance mass spectrometry platforms (Q-TOF and Orbitrap). We found that hpRP-LC consistently offered better resolution, reduced expression ratio compression, and a more MS-compatible workflow than OGE and consistently yielded more unique peptide/protein identifications and higher sequence coverage with better quantification. In this study, a total of 4463 potato proteins were identified, of which 46 showed differential expressions during potato tuber cold storage. Several key proteins important in controlling starch-sugar conversion, which leads to cold-induced sweetening, as well as other proteins that are potentially involved in this process, were identified. Our results suggest that the hpRP-RP shotgun approach is a feasible and practical workflow for discovering potential protein candidates in plant proteomic analysis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carbohydrate Metabolism
  • Chromatography, Liquid / methods*
  • Cryopreservation
  • Food Handling / methods
  • Food Preservation / methods
  • Hydrogen-Ion Concentration
  • Mass Spectrometry / methods*
  • Peptides / chemistry
  • Plant Proteins / chemistry
  • Proteomics / methods*
  • Reproducibility of Results
  • Solanum tuberosum / metabolism
  • Tandem Mass Spectrometry / methods

Substances

  • Peptides
  • Plant Proteins