Proteome-wide identification of ubiquitylation sites by conjugation of engineered lysine-less ubiquitin

Kiyotaka Oshikawa, Masaki Matsumoto, Koji Oyamada, Keiichi I. Nakayama

研究成果: ジャーナルへの寄稿記事

27 引用 (Scopus)

抄録

Ubiquitin conjugation (ubiquitylation) plays important roles not only in protein degradation but also in many other cellular functions. However, the sites of proteins that are targeted for such modification have remained poorly characterized at the proteomic level. We have now developed a method for the efficient identification of ubiquitylation sites in target proteins with the use of an engineered form of ubiquitin (K0-Ub), in which all seven lysine residues are replaced with arginine. K0-Ub is covalently attached to lysine residues of target proteins via an isopeptide bond, but further formation of a polyubiquitin chain does not occur on K0-Ub. We identified a total of 1392 ubiquitylation sites of 794 proteins from HEK293T cells. Profiling of ubiquitylation sites indicated that the sequences surrounding lysine residues targeted for ubiquitin conjugation do not share a common motif or structural feature. Furthermore, we identified a critical ubiquitylation site of the cyclin-dependent kinase inhibitor p27 Kip1. Mutation of this site thus inhibited ubiquitylation of and stabilized p27 Kip1, suggesting that this lysine residue is the target site of p27 Kip1 for ubiquitin conjugation in vivo. In conclusion, our method based on K0-Ub is a powerful tool for proteome-wide identification of ubiquitylation sites of target proteins.

元の言語英語
ページ(範囲)796-807
ページ数12
ジャーナルJournal of Proteome Research
11
発行部数2
DOI
出版物ステータス出版済み - 2 3 2012

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Ubiquitination
Proteome
Ubiquitin
Lysine
Proteins
Polyubiquitin
Cyclin-Dependent Kinase Inhibitor p27
Arginine
Proteomics
Proteolysis
Degradation
Mutation

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Chemistry(all)

これを引用

Proteome-wide identification of ubiquitylation sites by conjugation of engineered lysine-less ubiquitin. / Oshikawa, Kiyotaka; Matsumoto, Masaki; Oyamada, Koji; Nakayama, Keiichi I.

:: Journal of Proteome Research, 巻 11, 番号 2, 03.02.2012, p. 796-807.

研究成果: ジャーナルへの寄稿記事

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