Abstract：
Connexins are polytopic domain membrane proteins that form hexameric hemichannels (HCs) which can assemble into gap junction channels (GJCs) at the interface of two neighboring cells. The HCs may be involved in ion and small-molecule transport across the cellular plasma membrane in response to various stimuli. Despite their importance, relatively few structures of connexin HCs are available to date, compared to the structures of the GJCs. Here, we describe a protocol for expression, purification, and nanodisc reconstitution of connexin-43 (Cx43) HCs, which we have recently structurally characterized using cryo-EM analysis. Application of similar protocols to other connexin family members will lead to breakthroughs in the understanding of the structure and function of connexin HCs.
摘要：
连接蛋白是多位结构域膜蛋白,其形成六聚体半通道(HC),其可以在两个相邻细胞的界面处组装成间隙连接通道(GJC)。响应于各种刺激,HC可以参与穿过细胞质膜的离子和小分子转运。尽管它们很重要,到目前为止，连接蛋白HC的结构相对较少，与GJC的结构相比。这里,我们描述了一个表达的协议，净化,连接蛋白-43（Cx43）HC的纳米圆盘重建，我们最近使用低温EM分析进行了结构表征。将类似的方案应用于其他连接蛋白家族成员将导致对连接蛋白HC的结构和功能的理解的突破。
