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bioRxiv · 10.1101/2023.05.23.542033

Structural Visualization of HECT-E3 Ufd4 accepting and transferring Ubiquitin to Form K29/K48-branched Polyubiquitination on N-degron

Mao, J.·Ai, H.·Wu, X.·Zheng, Q.·Cai, H.·Liang, L.·Tong, Z.·Pan, M.·LIU, L.
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Abstract

The K29/K48-linked ubiquitination generated by the cooperative catalysis of E3 ligase Ufd4 and Ubr1 is an enhanced protein degradation signal, in which Ufd4 is responsible for introducing K29-linked ubiquitination to K48-linked ubiquitin chains to augment polyubiquitination. How HECT-E3 ligase Ufd4 mediates the ubiquitination event remains unclear. Here, we biochemically determine that Ufd4 preferentially catalyses K29-linked ubiquitination on K48-linked ubiquitin chains to generate K29/K48-branched ubiquitin chains and capture structural snapshots of Ub transfer cascades for Ufd4-mediated ubiquitination. The N-terminal ARM region and HECT domain C-lobe of Ufd4 are identified and characterized as key structural elements that together recruit K48-linked diUb and orient Lys29 of its proximal Ub to the active cysteine of Ufd4 for K29-linked branched ubiquitination. These structures not only provide mechanistic insights into the architecture of the Ufd4 complex but also provide structural visualization of branched ubiquitin chain formation by a HECT-type E3 ligase.

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Mao, J., Ai, H., Wu, X., Zheng, Q., Cai, H., Liang, L., Tong, Z., Pan, M., LIU, L.. 2023-05-24. Structural Visualization of HECT-E3 Ufd4 accepting and transferring Ubiquitin to Form K29/K48-branched Polyubiquitination on N-degron. https://doi.org/10.1101/2023.05.23.542033

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