Cai Ruiqi, Tang Jingfeng, Chen Xing-Zhen
Department of Physiology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB T6G 2H7, Canada.
National "111" Center for Cellular Regulation and Molecular Pharmaceutics, Hubei University of Technology, Wuhan, Hubei 430086, China.
iScience. 2021 Nov 1;24(12):103395. doi: 10.1016/j.isci.2021.103395. eCollection 2021 Dec 17.
Recently identified proton-activated chloride channel (PAC) contains two transmembrane helices (S1-S2) and is involved in lysosome function, hypoxia adaption, stroke, and carcinogenesis. Although a PAC structure was recently resolved, its gating and activation mechanisms remained largely unknown. By the two-electrode voltage clamp electrophysiology in oocytes, we found that the hydrophobicity of site 304 at fenestrations, but not that of neighbor sites, is important for maintaining PAC at a closed state at pH 7.5. When activated at acidic pH, PAC activity significantly increased with the hydrophilicity of site 307 within S2, but not with that of neighbor sites, suggesting that 307 acts as an activation gate. We identified six conserved protonatable residues critical for proton-induced activation, consistent with structural studies. Our study depicted a scheme in which proton binding induces conformational changes from the W304-controlled closed state at fenestrations to an activated state controlled by activation gate I307 in helix S2.
最近发现的质子激活氯离子通道（PAC）包含两个跨膜螺旋（S1-S2），并参与溶酶体功能、缺氧适应、中风和致癌作用。尽管最近解析了PAC的结构，但其门控和激活机制在很大程度上仍然未知。通过在卵母细胞中进行双电极电压钳电生理学实验，我们发现窗孔处304位点的疏水性而非相邻位点的疏水性对于在pH 7.5时将PAC维持在关闭状态很重要。当在酸性pH下激活时，PAC活性随着S2内307位点的亲水性显著增加，而不是随着相邻位点的亲水性增加，这表明307作为激活门。我们确定了六个对质子诱导的激活至关重要的保守可质子化残基，这与结构研究一致。我们的研究描绘了一个方案，其中质子结合诱导构象变化，从窗孔处由W304控制的关闭状态转变为由螺旋S2中的激活门I307控制的激活状态。
