Subunit connectivity, assembly determinants and architecture of the yeast exocyst complex.
Subunit connectivity, assembly determinants and architecture of the yeast exocyst complex.
复制标题
DOI：
10.1038/nsmb.3146
复制
发表时间：
2016-01
影响因子：
16.8
通讯作者：
Munson M
中科院分区：
文献类型：
--
作者：
Heider MR;Gu M;Duffy CM;Mirza AM;Marcotte LL;Walls AC;Farrall N;Hakhverdyan Z;Field MC;Rout MP;Frost A;Munson M
胞吐是一种异八聚体复合物，被认为是胞吐作用的束缚复合物，尽管在分子水平上对它的了解仍然很少。在这里，我们纯化的内源性外囊从酿酒酵母，并表明，纯化的复合物是稳定的，由所有八个亚基具有相等的化学计量。使用生化和生长素诱导的酵母降解实验的组合，我们映射的亚基连接，确定了两个稳定的四亚基模块内的八聚体，并证明了几个已知的外囊结合伙伴是不必要的外囊组装和稳定性。此外，我们可视化的酵母复合物的结构，使用负染色电子显微镜，我们的研究结果表明，外囊主要存在作为一个稳定的，八聚体复合物与一个细长的架构，这表明亚基是连续的螺旋束包装在一起成一束长杆。
The exocyst is a hetero-octameric complex proposed to serve as the tethering complex for exocytosis, although it remains poorly understood at the molecular level. Here, we purified endogenous exocyst from Saccharomyces cerevisiae, and show that the purified complexes are stable and consist of all eight subunits with equal stoichiometry. Using a combination of biochemical and auxin-induced degradation experiments in yeast, we mapped the subunit connectivity, identified two stable four-subunit modules within the octamer, and demonstrated that several known exocyst binding partners are not necessary for exocyst assembly and stability. Furthermore, we visualized the structure of the yeast complex using negative stain electron microscopy; our results indicate that exocyst exists predominantly as a stable, octameric complex with an elongated architecture that suggests the subunits are contiguous helical bundles packed together into a bundle of long rods.