Cell-Wide Survey of Amide-Bonded Lysine Modifications by Using Deacetylase CobB

Springer Science and Business Media LLC - Tập 21 Số 1 - 2019
Ying Wei1,2, Weiwei Yang1,2, Qijun Wang3, Peng-Cheng Lin4, Jin Zhao1,2, Wei Xu1,2, Shimin Zhao1,4,2, Xinlei He1,2
1Institutes of Biomedical Sciences, Obstetrics & Gynecology Hospital of Fudan University, State Key Lab of Genetic Engineering and School of Life Sciences, Shanghai, People’s Republic of China
2NHC Key Lab. of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Fudan University, Shanghai, People's Republic of China
3Shanghai Institute of Immunology, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China
4Key Laboratory for Tibet Plateau Phytochemistry of Qinghai Province, College of Pharmacy, Qinghai University for Nationalities, Xining, People’s Republic of China

Tóm tắt

AbstractBackgroundLysine post-translational modifications are important regulators of protein function. Proteomic and biochemical approaches have resulted in identification of several lysine modifications, including acetylation, crotonylation, and succinylation. Here, we developed an approach for surveying amide-bonded lysine modifications in the proteome of human tissues/cells based on the observation that many lysine modifications are amide-bonded and that theSalmonella entericadeacetylase, CobB, is an amidase.ResultsAfter the proteome of human tissues/cells was denatured and the non-covalently bonded metabolites were removed by acetone washes, and the amide-bonded modifiers were released by CobB and analyzed using liquid- and/or gas chromatography/mass spectrometry metabolomic analysis. This protocol, which required 3–4 days for completion, was used to qualitatively identify more than 40 documented and unreported lysine modifications from the human proteome and to quantitatively analyze dynamic changes in targeted amide-bonded lysine modifications.ConclusionsWe developed a method that was capable of monitoring and quantifying amide-bonded lysine modifications in cells of different origins.

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Tập 21 Số 1

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