Ministry of Education Key Laboratory of Protein Science, Tsinghua-Peking Joint Center for Life Sciences, Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Department of Anesthesiology, Shanghai First Maternity and Infant Hospital, Tongji University School of Medicine, Shanghai 201204, China.
Sci Adv. 2020 Aug 12;6(33):eabb4747. doi: 10.1126/sciadv.abb4747. eCollection 2020 Aug.
CLC family proteins translocate chloride ions across cell membranes to maintain the membrane potential, regulate the transepithelial Cl transport, and control the intravesicular pH among different organelles. CLC-7/Ostm1 is an electrogenic Cl/H antiporter that mainly resides in lysosomes and osteoclast ruffled membranes. Mutations in human CLC-7/Ostm1 lead to lysosomal storage disorders and severe osteopetrosis. Here, we present the cryo-electron microscopy (cryo-EM) structure of the human CLC-7/Ostm1 complex and reveal that the highly glycosylated Ostm1 functions like a lid positioned above CLC-7 and interacts extensively with CLC-7 within the membrane. Our complex structure reveals a functionally crucial domain interface between the amino terminus, TMD, and CBS domains of CLC-7. Structural analyses and electrophysiology studies suggest that the domain interaction interfaces affect the slow gating kinetics of CLC-7/Ostm1. Thus, our study deepens understanding of CLC-7/Ostm1 transporter and provides insights into the molecular basis of the disease-related mutations.
CLC 家族蛋白将氯离子跨细胞膜转运，以维持膜电位、调节跨上皮 Cl 转运，并控制不同细胞器之间的囊内 pH 值。CLC-7/Ostm1 是一种电活性的 Cl-/H+反向转运体，主要位于溶酶体和破骨细胞皱褶膜中。人类 CLC-7/Ostm1 的突变导致溶酶体贮积症和严重的骨质增生。在这里，我们呈现了人源 CLC-7/Ostm1 复合物的冷冻电镜（cryo-EM）结构，并揭示了高度糖基化的 Ostm1 像一个盖子位于 CLC-7 上方，并在膜内与 CLC-7 广泛相互作用。我们的复合物结构揭示了 CLC-7 的氨基末端、TMD 和 CBS 结构域之间的功能关键的结构域界面。结构分析和电生理学研究表明，结构域相互作用界面影响 CLC-7/Ostm1 的缓慢门控动力学。因此，我们的研究加深了对 CLC-7/Ostm1 转运蛋白的理解，并为疾病相关突变的分子基础提供了线索。
