Hutchison James M, Capone Ricardo, Luu Dustin D, Hadziselimovic Arina, Van Horn Wade D, Sanders Charles R
Chemical and Physical Biology Graduate Program, Vanderbilt University, Nashville, TN, 37240 USA.
Center for Structural Biology, Vanderbilt University, Nashville, TN, 37240 USA.
bioRxiv. 2021 Feb 19:2021.02.18.431684. doi: 10.1101/2021.02.18.431684.
SARS-CoV-2 envelope protein (S2-E) is a conserved membrane protein that is essential to coronavirus assembly and budding. Here, we describe the recombinant expression and purification of S2-E into amphipol-class amphipathic polymer solutions. The physical properties of amphipols underpin their ability to solubilize and stabilize membrane proteins without disrupting membranes. Amphipol delivery of S2-E to pre-formed planar bilayers results in spontaneous membrane integration and formation of viroporin ion channels. Amphipol delivery of the S2-E protein to human cells results in membrane integration followed by retrograde trafficking to a location adjacent to the endoplasmic reticulum-to-Golgi intermediate compartment (ERGIC) and the Golgi, which are the sites of coronavirus replication. Delivery of S2-E to cells enables both chemical biological approaches for future studies of SARS-CoV-2 pathogenesis and development of "Trojan Horse" anti-viral therapies. This work also establishes a paradigm for amphipol-mediated delivery of membrane proteins to cells.
严重急性呼吸综合征冠状病毒2包膜蛋白（S2-E）是一种保守的膜蛋白，对冠状病毒的组装和出芽至关重要。在此，我们描述了S2-E在两亲性聚合物溶液中的重组表达和纯化。两亲分子的物理性质决定了它们在不破坏膜的情况下溶解和稳定膜蛋白的能力。将S2-E通过两亲分子递送至预先形成的平面双层膜会导致其自发整合到膜中并形成病毒孔蛋白离子通道。将S2-E蛋白通过两亲分子递送至人类细胞会导致其整合到膜中，随后逆行运输至与内质网-高尔基体中间区室（ERGIC）和高尔基体相邻的位置，而这些位置正是冠状病毒复制的场所。将S2-E递送至细胞为未来研究严重急性呼吸综合征冠状病毒2发病机制的化学生物学方法以及“特洛伊木马”抗病毒疗法的开发提供了可能。这项工作还建立了一种通过两亲分子介导将膜蛋白递送至细胞的范例。
