Determinants of E2-ubiquitin conjugate recognition by RBR E3 ligases
Determinants of E2-ubiquitin conjugate recognition by RBR E3 ligases
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DOI：
10.1038/s41598-017-18513-5
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发表时间：
2018-01-08
影响因子：
4.6
通讯作者：
Rittinger, Katrin
中科院分区：
文献类型：
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作者：
Martino, Luigi;Brown, Nicholas R.;Rittinger, Katrin
环间环（RBR）泛素连接酶与多种E2酶一起工作，并通过E3-泛素硫酯中间体发挥作用。RBR模块包括三个结构域，RING 1、IBR和RING 2，它们协作以将泛素从类似于由RING 1识别的Ub缀合物的E2转移到RING 2中的催化半胱氨酸上，并最终在多步反应中转移到底物上。最近的研究表明，RING 1结构域结合E2类似于Ub共轭物在一个开放的构象，以抑制泛素转移到赖氨酸残基，并促进E3硫酯中间体的形成。然而，E2的性质如何影响泛素转移过程目前尚不清楚。我们在这里报告的RBR/E2-共轭识别步骤的详细表征表明，这种机制取决于E2酶的性质和UbcH 5和UbcH 7之间的不同。在类似于Ub的UbcH 5的情况下，与泛素的相互作用是稳定转移复合物所必需的，而类似于Ub的UbcH 7的识别主要由E2-RING 1接触驱动。此外，我们的分析表明，RBRs，在隔离和复杂的泛素负载E2，是动态的物种，其内在的灵活性可能是其催化机制的一个关键方面。
RING-between-RING (RBR) ubiquitin ligases work with multiple E2 enzymes and function through an E3-ubiquitin thioester intermediate. The RBR module comprises three domains, RING1, IBR and RING2 that collaborate to transfer ubiquitin from the E2 similar to Ub conjugate, recognised by RING1, onto a catalytic cysteine in RING2 and finally onto the substrate in a multi-step reaction. Recent studies have shown that RING1 domains bind E2 similar to Ub conjugates in an open conformation to supress ubiquitin transfer onto lysine residues and promote formation of the E3 thioester intermediate. However, how the nature of the E2 influences the ubiquitin transfer process is currently unclear. We report here a detailed characterization of the RBR/E2-conjugate recognition step that indicates that this mechanism depends on the nature of the E2 enzyme and differs between UbcH5 and UbcH7. In the case of UbcH5 similar to Ub an interaction with ubiquitin is necessary to stabilize the transfer complex while recognition of UbcH7 similar to Ub is driven primarily by E2-RING1 contacts. Furthermore our analysis suggests that RBRs, in isolation and in complex with ubiquitin-loaded E2s, are dynamic species and that their intrinsic flexibility might be a key aspect of their catalytic mechanism.