Binding of Substrates to the Central Pore of the Vps4 ATPase Is Autoinhibited by the Microtubule Interacting and Trafficking (MIT) Domain and Activated by MIT Interacting Motifs (MIMs)
Binding of Substrates to the Central Pore of the Vps4 ATPase Is Autoinhibited by the Microtubule Interacting and Trafficking (MIT) Domain and Activated by MIT Interacting Motifs (MIMs)
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DOI：
10.1074/jbc.m115.642355
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发表时间：
2015-05-22
影响因子：
4.8
通讯作者：
Hill, Christopher P.
中科院分区：
文献类型：
--
作者：
Han, Han;Monroe, Nicole;Hill, Christopher P.
转运所需的内体分选复合物（ESCRT）途径驱动多个细胞途径内的反向拓扑结构膜裂变事件，包括胞质分裂、多泡体生物发生、质膜修复、核膜囊泡形成和HIV出芽。AAA ATP酶Vps 4在裂变前不久被募集到膜颈，在那里它催化ESCRT-III晶格的分解。N-末端Vps 4微管相互作用和运输（MIT）结构域最初结合ESCRT-III亚基的C-末端MIT相互作用基序（MIM），但目前尚不清楚酶如何重塑这些底物以响应ATP水解。在这里，我们报告的定量结合研究表明，从螺旋5的Vps 2 p亚基的ESCRT-III的残基结合到一个不对称的Vps 4p六聚体的中心孔的方式，这是依赖于灵活的核苷酸类似物，可以模拟在ATP水解循环中的多个状态的存在。我们还发现，基板的接合是自抑制的Vps 4p MIT结构域，这种抑制被解除的结合，无论是1型或2型MIM元件，结合Vps 4p MIT结构域通过不同的接口。这些观察结果支持Vps 4底物最初通过MIM-MIT相互作用被募集的模型，所述MIM-MIT相互作用激活Vps 4中心孔以接合底物并产生力，从而触发ESCRT-III拆卸。
The endosomal sorting complexes required for transport (ESCRT) pathway drives reverse topology membrane fission events within multiple cellular pathways, including cytokinesis, multivesicular body biogenesis, repair of the plasma membrane, nuclear membrane vesicle formation, and HIV budding. The AAA ATPase Vps4 is recruited to membrane necks shortly before fission, where it catalyzes disassembly of the ESCRT-III lattice. The N-terminal Vps4 microtubule-interacting and trafficking (MIT) domains initially bind the C-terminal MIT-interacting motifs (MIMs) of ESCRT-III subunits, but it is unclear how the enzyme then remodels these substrates in response to ATP hydrolysis. Here, we report quantitative binding studies that demonstrate that residues from helix 5 of the Vps2p subunit of ESCRT-III bind to the central pore of an asymmetric Vps4p hexamer in a manner that is dependent upon the presence of flexible nucleotide analogs that can mimic multiple states in the ATP hydrolysis cycle. We also find that substrate engagement is autoinhibited by the Vps4p MIT domain and that this inhibition is relieved by binding of either Type 1 or Type 2 MIM elements, which bind the Vps4p MIT domain through different interfaces. These observations support the model that Vps4 substrates are initially recruited by an MIM-MIT interaction that activates the Vps4 central pore to engage substrates and generate force, thereby triggering ESCRT-III disassembly.