On the Possibility That Bond Strain Is the Mechanism of RING E3 Activation in the E2-Catalyzed Ubiquitination Reaction
On the Possibility That Bond Strain Is the Mechanism of RING E3 Activation in the E2-Catalyzed Ubiquitination Reaction
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DOI：
10.1021/acs.jcim.2c00423
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发表时间：
2022-06
影响因子：
5.6
通讯作者：
Jay-Anne K. Johnson;I. Sumner
中科院分区：
文献类型：
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作者：
Jay-Anne K. Johnson;I. Sumner
泛素化是一种翻译后修饰，其中小蛋白泛素(Ub)与靶蛋白上的赖氨酸共价结合。泛素化可以作为包括蛋白质降解在内的几种调控途径的信号。泛素化是由三种酶催化的一系列反应：泛素激活酶(E1)、泛素结合酶(E2)和泛素连接酶(E3)。E2酶直接催化Ub向靶蛋白─的转移，E3环提高了效率。在其转移之前，Ub通过硫酯键共价连接到E2，并且Ub∼E2共轭物与环E3形成四元络合物。据推测，E3环通过将E2∼Ub共轭物置于“闭合”位置而提高了泛素化的催化效率，从而拉紧并削弱了硫酯键。我们通过分析硫酯上的应变来验证这一假设，同时对开放和关闭的E2∼Ub/E3复合物进行了分子动力学模拟。我们的数据表明，当E2∼Ub共轭处于闭合位置时，硫酯处于应变状态。我们还表明，应变量与E3环引起的实验速率增强是一致的。最后，我们的模拟表明，闭合构型增加了E2∼Ub活性中心的关键氢键布居。这与另一个假设是一致的，即环E3通过预先组织底物来提高反应速度。
Ubiquitination is a type of post-translational modification wherein the small protein ubiquitin (Ub) is covalently bound to a lysine on a target protein. Ubiquitination can signal for several regulatory pathways including protein degradation. Ubiquitination occurs by a series of reactions catalyzed by three types of enzymes: ubiquitin activating enzymes, E1; ubiquitin conjugating enzymes, E2; and ubiquitin ligases, E3. E2 enzymes directly catalyze the transfer of Ub to the target protein─the RING E3 improves the efficiency. Prior to its transfer, Ub is covalently linked to the E2 via a thioester bond and the Ub∼E2 conjugate forms a quaternary complex with the RING E3. It is hypothesized that the RING E3 improves the catalytic efficiency of ubiquitination by placing the E2∼Ub conjugate in a "closed" position, which tensions and weakens the thioester bond. We interrogate this hypothesis by analyzing the strain on the thioester during molecular dynamics simulations of both open and closed E2∼Ub/E3 complexes. Our data indicate that the thioester is strained when the E2∼Ub conjugate is in the closed position. We also show that the amount of strain is consistent with the experimental rate enhancement caused by the RING E3. Finally, our simulations show that the closed configuration increases the populations of key hydrogen bonds in the E2∼Ub active site. This is consistent with another hypothesis stating that the RING E3 enhances reaction rates by preorganizing the substrates.