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泛素结合酶J2家族介导的泛素化反应中化学选择性的决定因素

Determinants of chemoselectivity in ubiquitination by the J2 family of ubiquitin-conjugating enzymes.

作者信息

Swarnkar Anuruti, Leidner Florian, Rout Ashok K, Ainatzi Sofia, Schmidt Claudia C, Becker Stefan, Urlaub Henning, Griesinger Christian, Grubmüller Helmut, Stein Alexander

机构信息

Research Group Membrane Protein Biochemistry, Max Planck Institute for Multidisciplinary Sciences, Am Fassberg 11, 37077, Göttingen, Germany.

Department of Theoretical and Computational Biophysics, Max Planck Institute for Multidisciplinary Sciences, Am Fassberg 11, 37077, Göttingen, Germany.

出版信息

EMBO J. 2024 Dec;43(24):6705-6739. doi: 10.1038/s44318-024-00301-3. Epub 2024 Nov 12.

DOI:10.1038/s44318-024-00301-3
PMID:39533056
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11649903/
Abstract

Ubiquitin-conjugating enzymes (E2) play a crucial role in the attachment of ubiquitin to proteins. Together with ubiquitin ligases (E3), they catalyze the transfer of ubiquitin (Ub) onto lysines with high chemoselectivity. A subfamily of E2s, including yeast Ubc6 and human Ube2J2, also mediates noncanonical modification of serines, but the structural determinants for this chemical versatility remain unknown. Using a combination of X-ray crystallography, molecular dynamics (MD) simulations, and reconstitution approaches, we have uncovered a two-layered mechanism that underlies this unique reactivity. A rearrangement of the Ubc6/Ube2J2 active site enhances the reactivity of the E2-Ub thioester, facilitating attack by weaker nucleophiles. Moreover, a conserved histidine in Ubc6/Ube2J2 activates a substrate serine by general base catalysis. Binding of RING-type E3 ligases further increases the serine selectivity inherent to Ubc6/Ube2J2, via an allosteric mechanism that requires specific positioning of the ubiquitin tail at the E2 active site. Our results elucidate how subtle structural modifications to the highly conserved E2 fold yield distinct enzymatic activity.

摘要

泛素结合酶（E2）在将泛素连接到蛋白质的过程中起着关键作用。它们与泛素连接酶（E3）一起，以高化学选择性催化泛素（Ub）转移到赖氨酸上。包括酵母Ubc6和人类Ube2J2在内的一个E2亚家族，也介导丝氨酸的非经典修饰，但这种化学多功能性的结构决定因素仍然未知。通过结合X射线晶体学、分子动力学（MD）模拟和重组方法，我们发现了一种两层机制，该机制是这种独特反应性的基础。Ubc6/Ube2J2活性位点的重排增强了E2-Ub硫酯的反应性，促进了较弱亲核试剂的攻击。此外，Ubc6/Ube2J2中一个保守的组氨酸通过一般碱催化激活底物丝氨酸。RING型E3连接酶的结合通过一种变构机制进一步提高了Ubc6/Ube2J2固有的丝氨酸选择性，该变构机制需要泛素尾巴在E2活性位点的特定定位。我们的结果阐明了对高度保守的E2折叠进行细微的结构修饰如何产生不同的酶活性。

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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/7cd2/11649903/87eded443d19/44318_2024_301_Fig8_ESM.jpg
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