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In Vitro Analysis of E3 Ubiquitin Ligase Function
06:06

In Vitro Analysis of E3 Ubiquitin Ligase Function

Published on: May 14, 2021

Activation of the E3 ubiquitin ligase Itch through a phosphorylation-induced conformational change

Ewen Gallagher1, Min Gao, Yun-Cai Liu

  • 1Laboratory of Gene Regulation and Signal Transduction, Department of Pharmacology, School of Medicine, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0723, USA.

Proceedings of the National Academy of Sciences of the United States of America
|February 1, 2006
PubMed

The E3 ubiquitin (Ub) ligase Itch is a critical regulator of T helper 2 (Th2) cytokine production through its ability to induce Ub-dependent JunB degradation. After T cell receptor engagement, Itch undergoes JNK1-mediated phosphorylation that greatly enhances its enzymatic activity. To investigate how phosphorylation activates an E3 Ub ligase we have identified the JNK1 phosphorylation sites within Itch as S199, S232, and T222, which are located within a Pro-rich region. Phosphorylation of these sites is necessary and sufficient for disrupting an inhibitory interaction between the WW domain of Itch and its catalytic HECT (Homologous to E6-AP C Terminus) domain and induces a conformational change that greatly enhances the catalytic activity of Itch, a HECT E3 ligase found to be directly activated upon its phosphorylation.

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