Journal
JOURNAL OF PROTEOME RESEARCH
Volume 10, Issue 10, Pages 4647-4660Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr200455s
Keywords
multiple dimensional separation; OffGel; high pH RP; LC-MS/MS; potato tubers; quantitative proteomics; cold storage; iTRAQ; Q-TOF; shotgun
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Funding
- ARS CRIS [1907-21000-024-00D]
- NSF [DBI-0606596]
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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 degrees 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.
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