Structure of E3 ligase E6AP with a proteasome-binding site provided by substrate receptor hRpn10
Structure of E3 ligase E6AP with a proteasome-binding site provided by substrate receptor hRpn10
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DOI：
10.1038/s41467-020-15073-7
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发表时间：
2020-03-10
影响因子：
16.6
通讯作者：
Walters, Kylie J.
中科院分区：
文献类型：
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作者：
Buel, Gwen R.;Chen, Xiang;Walters, Kylie J.
通过蛋白酶体调节的蛋白质水解涉及类似于800种用于用泛素修饰底物的酶，包括类似于600种E3连接酶。我们在这里报告，E6 AP/UBE 3A是区别于其他E3连接酶，在底物受体hRpn 10/PSMD 4/S5 a贡献的蛋白酶体上有一个12 nM的结合位点。hRpn 10中的E6 AP结合结构域本身是无序的，并且先前未被表征，它锁定成明确的螺旋结构，与E6 AP阿苏尔形成分子间4螺旋束，这是E3所特有的。因此，我们将hRpn 10 AZUL结合结构域命名为RAZUL。我们进一步发现，在人类细胞中，基于CRISPR的基因编辑导致RAZUL的丢失导致蛋白酶体中E6 AP的丢失。此外，蛋白酶体相关的泛素减少后，E6 AP敲低或从蛋白酶体的位移，表明E6 AP泛素化底物或蛋白酶体。总之，我们的研究结果表明，E6 AP是一个特权的E3蛋白酶体，一个专用的，高亲和力的结合位点贡献的hRpn 10。
Regulated proteolysis by proteasomes involves similar to 800 enzymes for substrate modification with ubiquitin, including similar to 600 E3 ligases. We report here that E6AP/UBE3A is distinguished from other E3 ligases by having a 12 nM binding site at the proteasome contributed by substrate receptor hRpn10/PSMD4/S5a. Intrinsically disordered by itself, and previously uncharacterized, the E6AP-binding domain in hRpn10 locks into a well-defined helical structure to form an intermolecular 4-helix bundle with the E6AP AZUL, which is unique to this E3. We thus name the hRpn10 AZUL-binding domain RAZUL. We further find in human cells that loss of RAZUL by CRISPR-based gene editing leads to loss of E6AP at proteasomes. Moreover, proteasome-associated ubiquitin is reduced following E6AP knockdown or displacement from proteasomes, suggesting that E6AP ubiquitinates substrates at or for the proteasome. Altogether, our findings indicate E6AP to be a privileged E3 for the proteasome, with a dedicated, high affinity binding site contributed by hRpn10.