Aman Teresa K, Gordon Sharona E, Zagotta William N
From the Department of Physiology and Biophysics, University of Washington, Seattle, Washington 98195.
From the Department of Physiology and Biophysics, University of Washington, Seattle, Washington 98195
J Biol Chem. 2016 May 6;291(19):9939-47. doi: 10.1074/jbc.M116.723932. Epub 2016 Mar 11.
Cyclic nucleotide-gated (CNG) channels are expressed in rod photoreceptors and open in response to direct binding of cyclic nucleotides. We have previously shown that potentiation of CNGA1 channels by transition metals requires a histidine in the A' helix following the S6 transmembrane segment. Here, we used transition metal ion FRET and patch clamp fluorometry with a fluorescent, noncanonical amino acid (3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap)) to show that the potentiating transition metal Co(2+) binds in or near the A' helix. Adding high-affinity metal-binding sites to the membrane (stearoyl-nitrilotriacetic acid (C18-NTA)) increased potentiation for low Co(2+) concentrations, indicating that the membrane can coordinate metal ions with the A' helix. These results suggest that restraining the A' helix to the plasma membrane potentiates CNGA1 channel opening. Similar interactions between the A' helix and the plasma membrane may underlie regulation of structurally related hyperpolarization-activated cyclic nucleotide-gated (HCN) and voltage-gated potassium subfamily H (KCNH) channels by plasma membrane components.
环核苷酸门控（CNG）通道在视杆光感受器中表达，并在环核苷酸直接结合时开放。我们之前已经表明，过渡金属对CNGA1通道的增强作用需要S6跨膜段之后A'螺旋中的一个组氨酸。在这里，我们使用过渡金属离子荧光共振能量转移（FRET）和膜片钳荧光法结合一种荧光非天然氨基酸（3-（6-乙酰萘-2-基氨基）-2-氨基丙酸（Anap））来表明，增强作用的过渡金属Co(2+)结合在A'螺旋内或其附近。向膜中添加高亲和力金属结合位点（硬脂酰亚氨基三乙酸（C18-NTA））可增强低浓度Co(2+)的增强作用，表明膜可以将金属离子与A'螺旋配位。这些结果表明，将A'螺旋限制在质膜上可增强CNGA1通道的开放。A'螺旋与质膜之间的类似相互作用可能是质膜成分对结构相关的超极化激活环核苷酸门控（HCN）通道和电压门控钾亚家族H（KCNH）通道进行调节的基础。
