A new perspesective on ion conductance and structural dynamics of ion channels using 2D IR

使用 2D IR 对离子电导和离子通道结构动力学的新视角

• 批准号: 10222727
• 负责人: Francis Valiyaveetil
• 金额: $ 34.09万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10222727
• 关键词: Address; Arrhythmia; Binding Sites; Biological; Cell membrane; Charge; Collaborations; Communities; Data; Dependence; Development; Disease; Electrostatics; Epilepsy; Family; Frequencies; Human; Integral Membrane Protein; Ion Channel; Ion Transport; Ions; Isotope Labeling; Kinetics; Lead; Light; Measurement; Measures; Membrane; Membrane Proteins; Methodology; Modeling; Molecular Conformation; Motion; Mutation; Physiology; Play; Potassium Channel; Procedures; Proteins; Publications; Research; Resolution; Roentgen Rays; Role; Science; Series; Site; Spectrum Analysis; Structural Models; Structure; Techniques; Technology; Testing; Textbooks; Time; Variant; Vertebral column; Water; Work; X-Ray Crystallography; absorption; base; electric field; experimental study; fascinate; innovation; insight; ion dynamics; millisecond; molecular dynamics; patch clamp; two-dimensional; vibration; voltage

Abstract Patch clamp experiments performed since the 1970s have provided the timescales for the opening and closing of ion channels. X-ray crystallography over the past 2 decades has yielded high-resolution structures of ion channels. As yet, there are no direct experiments on channel dynamics or the effect of an applied voltage on ion channel structures. In this proposal, we connect dynamics to structure by leveraging new technological advances that enable two-dimensional infrared (2D IR) measurements on ion channels. The inherent time-resolution of 2D IR spectroscopy is a few picoseconds – much faster than the millisecond motions of ion channels. Structural resolution arises from couplings between the backbone carbonyl vibrations and electrostatic charges such as ions. Residue-specific to domain-specific structural resolution is obtained with isotope labeling made routine by semisynthesis procedures developed in the Valiyaveetil lab. And, 2D IR spectra can now be calculated very accurately from short molecular dynamics trajectories, enabling a one-to-one comparison between experiment and structure or proposed structural models. This combination of 2D IR, semisynthesis, and molecular dynamics simulations permit a new perspective on ion channel structural dynamics. In this proposal, we address two outstanding controversies in the potassium ion channel community. In Aims 1 and 2, we investigate a controversial new model for ion permeation through the selectivity filter of KcsA and NaK, called the “hard-knock” model. The hard-knock model appears to explain the X-ray data and all other existing measurements, even though it is fundamentally at odds with the original “knock-on” model found in textbooks. In Aim 3, we voltage- trigger the structural motions of the voltage sensing domain (VSD) of the KvAP channel, to investigate the hypothesis that the essential TM4 helix in the VSD undergoes a conformational and/or hydrational change during voltage gating, as previously proposed but never established by a direct structural or time-resolved measurement. These aims provide new scientific insights into outstanding problems in the ion channel community and establish a new technique for studying ion channel structural dynamics.

摘要