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  3. Two-dimensional gel electrophoretic detection of protein carbonyls derivatized with biotin-hydrazide

Two-dimensional gel electrophoretic detection of protein carbonyls derivatized with biotin-hydrazide

  • J Chromatogr B Analyt Technol Biomed Life Sci. 2016 Apr 15;1019:128-31. doi: 10.1016/j.jchromb.2015.11.003.
Jinzi Wu 1,  Xiaoting Luo 2,  Siqun Jing 3,  Liang-Jun Yan 4
Affiliations

Affiliations

  • 1 Department of Pharmaceutical Sciences, UNT System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, TX 76107, United States.
  • 2 Department of Pharmaceutical Sciences, UNT System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, TX 76107, United States; Department of Biochemistry and Molecular Biology, Gannan Medical University, Ganzhou, Jiangxi Province 341000, China.
  • 3 Department of Pharmaceutical Sciences, UNT System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, TX 76107, United States; College of Life Sciences and Technology, Xinjiang University, Urumqi, Xinjiang 830046, China.
  • 4 Department of Pharmaceutical Sciences, UNT System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, TX 76107, United States. Electronic address: liang-jun.yan@unthsc.edu.
PMID: 26590475 DOI: 10.1016/j.jchromb.2015.11.003
Abstract

Protein carbonyls are protein oxidation products that are often used to measure the magnitude of protein oxidative damage induced by reactive oxygen or reactive nitrogen species. Protein carbonyls have been found to be elevated during aging and in age-related diseases such as stroke, diabetes, and neurodegenerative diseases. In the present article, we provide detailed protocols for detection of mitochondrial protein carbonyls labeled with biotin-hydrazide followed by 2-dimensional isoelectric focusing (IEF)/SDS-PAGE and Western blotting probed with horse-radish peroxidase-conjugated streptavidin. The presented procedures can also be modified for detection of carbonylation of non-mitochondrial proteins.

Keywords

Biotin-hydrazide; Carbonylation; IEF/SDS-PAGE; Mitochondria; Protein carbonyls; Protocol; Streptavidin.

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