An Improved Isotope-coded Affinity Tag Technology for Thiol Redox Proteomics

DOI: 10.5584/jiomics.v2i1.83

Authors

  • Ning Zhu Department of Biology, Genetics Institute, University of Florida, Gainesville, FL 32610, USA
  • Mengmeng Zhu Department of Biology, Genetics Institute, University of Florida, Gainesville, FL 32610, USA
  • Shaojun Dai Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline-alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040, China
  • Ran Zheng Proteomics Facility, Interdisciplinary Center for Biotechnology Research, University of Florida, Gainesville, FL 32610, USA
  • Sixue Chen Proteomics Facility, Interdisciplinary Center for Biotechnology Research, University of Florida, Gainesville, FL 32610, USA

Abstract

Isotope coded affinity tag (ICAT) is a gel-free technology for quantitative proteomics. In ICAT procedure, strong cation exchange chromatography (SCX) using increased potassium chloride gradient is recommended for peptide fractionation. Here we report optimization of hydrophilic interaction chromatography (HILIC) as an alternative strategy for peptide fractionation of ICAT samples. HILIC exhibits high separation efficiency and does not require any downstream desalting steps. Compared to SCX based ICAT, integration of HILIC into the ICAT technology has resulted in high rates of protein identification, cysteine mapping, and quantification of cysteine-containing peptides. The improved technology has shown utility in thiol redox proteomics. Interestingly, results from HILIC ICAT and SCX ICAT are complementary. Implementation of both provides high coverage analysis of a complex proteome.

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Published

2012-05-31

Issue

Vol. 2 No. 1 (2012): Vol 2, No 1 (2012)

Section

Articles