Department of Biochemistry, Brandeis University, Waltham, MA 02453.
Department of Pediatrics, Loyola University Chicago Stritch School of Medicine, Maywood, IL 60153.
Proc Natl Acad Sci U S A. 2018 Dec 11;115(50):E11847-E11856. doi: 10.1073/pnas.1816189115. Epub 2018 Nov 26.
Using a de novo peptide inhibitor, Corza6 (C6), we demonstrate that the human voltage-gated proton channel (hHv1) is the main pathway for H efflux that allows capacitation in sperm and permits sustained reactive oxygen species (ROS) production in white blood cells (WBCs). C6 was identified by a phage-display strategy whereby ∼1 million novel peptides were fabricated on an inhibitor cysteine knot (ICK) scaffold and sorting on purified hHv1 protein. Two C6 peptides bind to each dimeric channel, one on the S3-S4 loop of each voltage sensor domain (VSD). Binding is cooperative with an equilibrium affinity () of ∼1 nM at -50 mV. As expected for a VSD-directed toxin, C6 inhibits by shifting hHv1 activation to more positive voltages, slowing opening and speeding closure, effects that diminish with membrane depolarization.
我们使用一种从头设计的肽抑制剂 Corza6（C6），证明了人类电压门控质子通道（hHv1）是 H 外流的主要途径，该途径允许精子获能，并允许白细胞（WBC）中持续产生活性氧（ROS）。C6 是通过噬菌体展示策略鉴定的，即用抑制剂半胱氨酸结（ICK）支架制造了大约 100 万个新肽，并在纯化的 hHv1 蛋白上进行了分拣。两个 C6 肽结合到每个二聚体通道上，一个结合在每个电压传感器域（VSD）的 S3-S4 环上。结合具有协同性，在-50 mV 时平衡亲和力（）约为 1 nM。正如针对 VSD 定向毒素的预期，C6 通过将 hHv1 的激活转移到更正的电压来抑制，从而减缓打开速度并加速关闭速度，这些作用随着膜去极化而减弱。
