Bimodular architecture of bacterial effector SAP05 drives ubiquitin-independent targeted protein degradation
Bimodular architecture of bacterial effector SAP05 drives ubiquitin-independent targeted protein degradation
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细菌效应器 SAP05 的双模块结构驱动不依赖于泛素的靶向蛋白质降解
DOI：
10.1101/2023.06.19.545293
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Liu Q
中科院分区：
文献类型：
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作者：
Liu Q
在真核生物中，靶向蛋白降解（TPD）通常依赖于泛素连接酶之间的一系列相互作用，所述泛素连接酶将泛素分子转移到底物，导致26 S蛋白酶体的降解。我们以前发现，细菌效应蛋白SAP 05介导的泛素非依赖性TPD。SAP 05通过与蛋白酶体泛素受体Rpn 10的A型血管性血友病因子（vWA）结构域和鳞状细胞启动子结合蛋白样（SPL）和加塔结合因子（加塔）转录因子（TF）的锌指（ZnF）结构域相互作用形成三元复合物。这导致TF通过26 S蛋白酶体的直接TPD。在这里，我们报告的晶体结构的SAP 05-Rpn 10 vWA复合物在2.17纳米分辨率和SAP 05-SPL 5 ZnFcomplex在2.20纳米分辨率。结构分析表明，SAP 05显示出一个显着的双模块结构，具有两个不同的非重叠表面，一个“环表面”，具有三个突出的环，形成与ZnF的静电相互作用，和一个“片表面”，具有两个β-片，环，和α-螺旋，建立极性相互作用与vWA。SAP 05与ZnF TF的结合涉及负责多个接触的单个氨基酸，而SAP 05与vWA的结合由于需要多个突变来打破相互作用而更稳定。此外，SAP 05复合物在26 S蛋白酶体上的定位指出了蛋白质降解的机制。总的来说，我们的研究结果表明，一个小的细菌双模蛋白可以绕过典型的泛素-蛋白酶体蛋白水解途径，使泛素独立的TPD在真核细胞。这些知识对TPD技术的创造具有重要的潜力。
In eukaryotes, targeted protein degradation (TPD) typically depends on a series of interactions among ubiquitin ligases that transfer ubiquitin molecules to substrates leading to degradation by the 26S proteasome. We previously identified that the bacterial effector protein SAP05 mediates ubiquitin-independent TPD. SAP05 forms a ternary complex via interactions with the von Willebrand Factor Type A (vWA) domain of the proteasomal ubiquitin receptor Rpn10 and the zinc-finger (ZnF) domains of the SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE (SPL) and GATA BINDING FACTOR (GATA) transcription factors (TFs). This leads to direct TPD of the TFs by the 26S proteasome. Here, we report the crystal structures of the SAP05–Rpn10vWAcomplex at 2.17 Å resolution and of the SAP05–SPL5ZnFcomplex at 2.20 Å resolution. Structural analyses revealed that SAP05 displays a remarkable bimodular architecture with two distinct nonoverlapping surfaces, a “loop surface” with three protruding loops that form electrostatic interactions with ZnF, and a “sheet surface” featuring two β-sheets, loops, and α-helices that establish polar interactions with vWA. SAP05 binding to ZnF TFs involves single amino acids responsible for multiple contacts, while SAP05 binding to vWA is more stable due to the necessity of multiple mutations to break the interaction. In addition, positioning of the SAP05 complex on the 26S proteasome points to a mechanism of protein degradation. Collectively, our findings demonstrate how a small bacterial bimodular protein can bypass the canonical ubiquitin–proteasome proteolysis pathway, enabling ubiquitin-independent TPD in eukaryotic cells. This knowledge holds significant potential for the creation of TPD technologies.