Improved mammalian retromer cryo-EM structures reveal a new assembly interface.
Improved mammalian retromer cryo-EM structures reveal a new assembly interface.
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DOI：
10.1016/j.jbc.2022.102523
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发表时间：
2022-11
影响因子：
4.8
通讯作者：
Jackson, Lauren P.
中科院分区：
文献类型：
--
作者：
Kendall, Amy K.;Chandra, Mintu;Xie, Boyang;Wan, William;Jackson, Lauren P.
Retromer（VPS 26/VPS 35/VPS 29亚基）与膜上的多个分选连接蛋白组装以介导跨膜蛋白货物的内体再循环。Retromer还参与其他细胞过程，包括线粒体稳态，营养感测，自噬和裂变事件。哺乳动物逆转录聚合物组装的机制仍然不确定，逆转录聚合物与多种分选连接蛋白蛋白结合，将货物分选到不同的目的地。已发表的结构证明哺乳动物retromer在体外形成低聚物，但几种结构解析不佳。我们在这里报告改进retromer低聚物结构，使用单粒子cryo-EM通过结合从倾斜标本收集的数据与数据处理的多个进步，包括使用3D起始模型增强自动化粒子拾取在RELION。我们使用了一个逆转录酶突变体（3 KE逆转录酶），打破VPS 35介导的接口，以确定一个新的组装界面的结构形成的VPS 26 A和VPS 35的N-末端。该界面揭示了N-末端VPS 26 A抑制蛋白鞍如何通过接合相邻的VPS 35 N-末端来连接逆转录链，所述N-末端在异源三聚体内观察到的充分表征的C-VPS 26/N-VPS 35相互作用的相对侧。新的相互作用界面表现出相当大的掩埋表面积（7000 μ 2），并进一步表明，后生动物retromer可能作为一个适应性的支架。
Retromer (VPS26/VPS35/VPS29 subunits) assembles with multiple sorting nexin proteins on membranes to mediate endosomal recycling of transmembrane protein cargoes. Retromer has been implicated in other cellular processes, including mitochondrial homeostasis, nutrient sensing, autophagy, and fission events. Mechanisms for mammalian retromer assembly remain undefined, and retromer engages multiple sorting nexin proteins to sort cargoes to different destinations. Published structures demonstrate mammalian retromer forms oligomers in vitro, but several structures were poorly resolved. We report here improved retromer oligomer structures using single-particle cryo-EM by combining data collected from tilted specimens with multiple advancements in data processing, including using a 3D starting model for enhanced automated particle picking in RELION. We used a retromer mutant (3KE retromer) that breaks VPS35-mediated interfaces to determine a structure of a new assembly interface formed by the VPS26A and VPS35 N-termini. The interface reveals how an N-terminal VPS26A arrestin saddle can link retromer chains by engaging a neighboring VPS35 N- terminus, on the opposite side from the well-characterized C-VPS26/N-VPS35 interaction observed within heterotrimers. The new interaction interface exhibits substantial buried surface area (∼7000 Å2) and further suggests that metazoan retromer may serve as an adaptable scaffold.
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影响因子：
64.5
作者：
Daniloski Z;Jordan TX;Wessels HH;Hoagland DA;Kasela S;Legut M;Maniatis S;Mimitou EP;Lu L;Geller E;Danziger O;Rosenberg BR;Phatnani H;Smibert P;Lappalainen T;tenOever BR;Sanjana NE
通讯作者：
Sanjana NE
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
5.3
作者：
Fjorback, Anja W.;Seaman, Matthew;Andersen, Olav M.
通讯作者：
Andersen, Olav M.
DOI：
10.1073/pnas.0503689102
发表时间：
2005-09-20
影响因子：
11.1
作者：
Andersen, OM;Reiche, J;Willnow, TE
通讯作者：
Willnow, TE
影响因子：
16.8
作者：
Collins, BM;Skinner, CF;Owen, DJ
通讯作者：
Owen, DJ