The Beatson Institute for Cancer Research, Garscube Estate, Switchback Road, Glasgow, G61 1BD, United Kingdom.
Nat Struct Mol Biol. 2012 Sep;19(9):876-83. doi: 10.1038/nsmb.2379. Epub 2012 Aug 14.
Certain RING ubiquitin ligases (E3s) dimerize to facilitate ubiquitin (Ub) transfer from ubiquitin-conjugating enzyme (E2) to substrate, but structural evidence on how this process promotes Ub transfer is lacking. Here we report the structure of the human dimeric RING domain from BIRC7 in complex with the E2 UbcH5B covalently linked to Ub (UbcH5B∼Ub). The structure reveals extensive noncovalent donor Ub interactions with UbcH5B and both subunits of the RING domain dimer that stabilize the globular body and C-terminal tail of Ub. Mutations that disrupt these noncovalent interactions or RING dimerization reduce UbcH5B∼Ub binding affinity and ubiquitination activity. Moreover, NMR analyses demonstrate that BIRC7 binding to UbcH5B∼Ub induces peak-shift perturbations in the donor Ub consistent with the crystallographically-observed Ub interactions. Our results provide structural insights into how dimeric RING E3s recruit E2∼Ub and optimize the donor Ub configuration for transfer.
某些环指泛素连接酶（E3s）二聚化以促进泛素（Ub）从泛素结合酶（E2）转移到底物，但缺乏关于该过程如何促进 Ub 转移的结构证据。在这里，我们报告了人源 BIRC7 二聚化环指结构域与共价连接 Ub 的 E2 UbcH5B（UbcH5B∼Ub）复合物的结构。该结构揭示了 UbcH5B 和 RING 结构域二聚体的两个亚基与供体 Ub 之间广泛的非共价相互作用，这些相互作用稳定了 Ub 的球状主体和 C 末端尾部。破坏这些非共价相互作用或 RING 二聚化的突变会降低 UbcH5B∼Ub 的结合亲和力和泛素化活性。此外，NMR 分析表明，BIRC7 与 UbcH5B∼Ub 的结合会导致供体 Ub 的峰移动扰动，与晶体学观察到的 Ub 相互作用一致。我们的结果提供了结构见解，说明了二聚化 RING E3 如何招募 E2∼Ub 并优化供体 Ub 的转移构型。
