Proton Conduction Pathways in Proton Channel Proteins

质子通道蛋白中的质子传导途径

• 批准号: 10039569
• 负责人: Huong Tran Kratochvil
• 金额: $ 9.72万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10039569
• 关键词: Acids; Address; Affect; Amantadine; Antiviral Agents; Binding; Bioenergetics; Biological Process; Biophysics; Cell membrane; Cells; Cellular Membrane; Charge; Crystallization; Crystallography; Diffusion; Disease; Drug resistance; Event; Glutamine; Hydration status; Hydrogen Bonding; Influenza; Influenza A virus; Ions; LGLA; Lead; Length; Lipid Bilayers; Measurement; Mediating; Membrane; Methods; Molecular; Molecular Conformation; Motion; Mutation; Pathway interactions; Permeability; Pharmaceutical Preparations; Phase; Play; Positioning Attribute; Process; Proteins; Protons; Replication-Associated Process; Resolution; Role; Signal Transduction; Site; Stream; Structure; Testing; Time; Vertebral column; Virus Replication; Water; Work; computer studies; deprotonation; design; infrared spectroscopy; molecular dynamics; mutant; novel; protonation; resistance mutation; transmission process; two-dimensional

PROJECT ABSTRACT: Proton channel proteins potentiate the flow of protons across cell membranes, and have evolved fine control over proton selectivity and conductivity to efficiently achieve their function, while maintaining cellular integrity. Through formation of dynamic proton conduction pathways which mimic the water wires observed in dilute acid for proton diffusion, protons move rapidly and selectively along a hydrogen-bonding network composed of confined water and ionizable sidechains scattered within the lumen of proton channel proteins. One way proton channels mediate proton conductivity is through guide water wires, which are stable lumenal waters organized by polar protein groups. Guide water wires are well-studied as they are observed in high-resolution crystal structures, but whether they are mobile or static and how their dynamics affects proton conductivity remains unclear. Another way to modulate proton selectivity and conductivity is through transient water wires, which are thought to form and dissipate to allow for proton flux through well-packed apolar segments. While transient water wires have been hypothesized in molecular dynamics (MD) simulations, they are fundamentally difficult to test experimentally. Finally, proton channels also use proton shuttle mechanisms of protonation and deprotonation through an ionizable sidechain, such as His, Glu, and Asp, to tune proton conductance, but it is unclear the extent these sidechains mediate pore solvation, and whether the proton shuttle mechanism leads to a net transit of water. This work will address these mechanisms by which proton channel proteins mediate proton flux: the (1) seemingly stable hydrogen-bonding networks of guide water wires and protein polar groups, (2) transient water wires, and (3) proton shuttles composed of ionizable sidechains. Through our proposed study of a natural proton channel, the influenza A matrix protein 2 (M2), and de novo designed proton channels, we will test the hypotheses that (1) guide and transient water wires within proton channel proteins confer their selectivity and dictate their capacity to conduct protons, and (2) proton shuttles are not only necessary in defining the conduction rates of these proton channels, but also play critical roles in modulating proton and water permeability. In Aim 1, we will examine whether guide water wires are mobile or static by multidimensional infrared spectroscopy on M2 proton channels and the disease-relative mutants. Our measurements in the presence and absence of drugs will allow us to determine how the dynamics of these networks affect proton conductance, and how they change with drug binding and resistance mutations, which is critical to identifying new antiviral strategies. In Aim 2, we test the hypothesis of transient water wires through the de novo design and characterization of novel proton channels with varying lengths of apolar regions. In the R00 phase (Aim 3), we examine how ionizable sidechains potentiate pore hydration and investigate whether protonation/deprotonation events lead to the cotranslocation of protons and water.

项目摘要： 质子通道蛋白增强了质子在细胞膜上的流动，并进化出精细的控制。 超质子选择性和传导性，以有效地实现其功能，同时保持细胞的完整性。 通过形成动态质子传导路径，模拟在稀酸中观察到的水线 对于质子扩散，质子沿着由以下组成的氢键网络快速和选择性地移动 分散在质子通道蛋白管腔内的承压水和可电离侧链。单程质子 中介质子电导的通道是通过导水管，这些导线是由稳定的腔水组织起来的。 通过极性蛋白质组。由于在高分辨率晶体中观察到了导水线，因此对它们进行了很好的研究 结构，但它们是运动的还是静态的，以及它们的动力学如何影响质子的传导性仍然存在 不清楚。另一种调节质子选择性和传导性的方法是通过瞬变水线，这是 被认为形成和消散，以允许质子通过紧密堆积的非极区段。而瞬息之水 在分子动力学(MD)模拟中已经假设了导线，它们从根本上很难测试 试验性的。最后，质子通道还使用质子化和去质子化的质子穿梭机制 通过可电离侧链，如His，Glu和Asp，来调节质子电导，但尚不清楚 这些侧链在多大程度上调节了孔溶剂化，以及质子穿梭机制是否导致了净迁移。 一桶水。这项工作将解决质子通道蛋白调节质子通量的这些机制： (1)看似稳定的导水线和蛋白质极性基团的氢键网络，(2)瞬时 水线，以及(3)由可电离侧链组成的质子航天飞机。 通过我们提议的天然质子通道、甲型流感病毒基质蛋白2(M2)和从头开始的研究 在设计的质子通道中，我们将检验(1)质子内部引导和瞬变水线的假设 通道蛋白赋予它们的选择性，并决定它们传导质子的能力，以及(2)质子航天飞机是 不仅在定义这些质子通道的传导率方面是必要的，而且在 调节质子和水的渗透性。在目标1中，我们将检查导水管是可移动的还是 在M2质子通道和疾病相关突变体上的多维红外光谱静态。我们的 在有无药物的情况下进行测量将使我们能够确定这些药物的动态 网络影响质子电导，以及它们如何随着药物结合和耐药性突变而变化，这是 对于确定新的抗病毒策略至关重要。在目标2中，我们通过对瞬变水线的假设进行检验 具有不同非极区长度的新型质子通道的从头设计和表征。在 R00相(目标3)，我们研究了可电离侧链如何增强孔隙水化，并调查了 质子化/去质子化事件导致质子和水的共移位。