CFTR基因的功能缺失突变导致缩短寿命的遗传病囊性纤维化(CF),而CFTR的过度活跃可能导致分泌性腹泻和多囊肾疾病。虽然已经开发了针对CFTR蛋白的有效药物用于治疗CF,由超激活CFTR引起的疾病进展甚微。这里,我们解决了CFTR与CFTRinh-172(Inh-172)复合的低温EM结构,具有有希望的效力和功效的CFTR门控抑制剂。我们发现Inh-172结合在CFTR的孔内,通过主要的疏水相互作用和盐桥与跨膜片段(TM)1、6、8、9和12的氨基酸残基相互作用。这些残基的取代降低了Inh-172的表观亲和力。抑制剂结合的结构揭示了TM1、8和12的细胞外段的重新定向,支持涉及结合后构象变化的变构调节机制。这种变构抑制机制很容易解释我们的观察结果,猪CFTR,它保留了所有参与Inh-172结合的氨基酸残基,对Inh-172的敏感性大大降低,并且CF药物ivacaftor改变了Inh-172的表观亲和力(即,VX-770),通过与还包含TM8的位点结合来增强CFTR的活性。
Loss-of-function mutations of the CFTR gene cause the life-shortening genetic disease cystic fibrosis (CF), whereas overactivity of CFTR may lead to secretory diarrhea and polycystic kidney disease. While effective drugs targeting the CFTR protein have been developed for the treatment of CF, little progress has been made for diseases caused by hyper-activated CFTR. Here, we solve the cryo-EM structure of CFTR in complex with CFTRinh-172 (Inh-172), a CFTR gating inhibitor with promising potency and efficacy. We find that Inh-172 binds inside the pore of CFTR, interacting with amino acid residues from transmembrane segments (TMs) 1, 6, 8, 9, and 12 through mostly hydrophobic interactions and a salt bridge. Substitution of these residues lowers the apparent affinity of Inh-172. The inhibitor-bound structure reveals re-orientations of the extracellular segment of TMs 1, 8, and 12, supporting an allosteric modulation mechanism involving post-binding conformational changes. This allosteric inhibitory mechanism readily explains our observations that pig CFTR, which preserves all the amino acid residues involved in Inh-172 binding, exhibits a much-reduced sensitivity to Inh-172 and that the apparent affinity of Inh-172 is altered by the CF drug ivacaftor (i.e., VX-770) which enhances CFTR\'s activity through binding to a site also comprising TM8.