Chemical biology of voltage-gated cation channels

电压门控阳离子通道的化学生物学

• 批准号: 10552311
• 负责人: Christopher A Ahern
• 金额: $ 53.51万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2028-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10552311
• 关键词: Adopted; Amplifiers; Arrhythmia; Binding; Binding Sites; Biological; Biology; Calcium Channel; Cardiac; Cations; Cell membrane; Cells; Chemicals; Chemistry; Clinical; Dissection; Environment; Epilepsy; Event; Foundations; Goals; Heart Rate; Heart failure; Human; Hypertension; Individual; Ion Channel; Ion Channel Gating; Ions; Light; Link; Long QT Syndrome; Mammalian Cell; Medicine; Membrane; Methods; Modification; Molecular; Muscle; Muscle Contraction; Neurons; Pain management; Pathway interactions; Peptides; Perception; Pharmaceutical Preparations; Phenylalanine; Phosphorylation; Physiological; Potassium Channel; Process; Protein Conformation; Proteins; Research; Resolution; Second Messenger Systems; Shapes; Signal Transduction; Site; Sodium Channel; Speed; Therapeutic; Tyrosine; Vertebral column; Voltage-Gated Potassium Channel; chemical binding; experimental study; extracellular; fighting; innovation; potassium ion; programs; protein structure; response; sensor; skeletal; training opportunity; voltage; voltage gated channel

Voltage-gated ion channels shape electrical signaling in the excitable cells of nerve and muscle. Sodium (NaV) and calcium channels (CaV) drive membrane depolarization and activate second messenger pathways via gated cellular entry of their namesake ions. In skeletal and cardiac cells, CaV channels trigger muscle contraction. Voltage-gated potassium channels (KV) allow the release of potassium ions from within the cell to drive membrane repolarization. In concert, these channels provide the molecular foundation for thought, perception, and contraction. High-resolution protein structures of human voltage-gated channels are now providing the first glimpses of the types of poses they may adopt in cellular environments. However, understanding the ultimate link between how these proteins look and how they support physiological mechanisms is a major challenge that will require innovative approaches. For one, transmembrane voltage is absent in a structural experiment thus depicting voltage-gated channels in an essentially non-physiological environment. We are therefore developing photochemical `stapling' approaches to covalently trap high-value protein conformations in live cell membranes prior to purification for structural determination. Further, we have begun to identify mechanisms of channel function by introducing modified chemistries at the peptide backbone in the transmembrane segments that form voltage-sensors and channel gates. In cellular settings, ion channels are also critical amplifiers of transduction pathways. During the fight-or-fight response, for instance, the near instantaneous phosphorylation of CaV1.2 channels results in faster and sustained channel opening, leading to a more forceful and rapid heart rate. Yet the absolute speed and complexity of the process is a challenge to experimentally parse individual molecular events that result in channel gating modifications. We describe newly validated methods that enable light controlled, site-specific phosphorylation, for the careful deconstruction and identification of key steps and players is this process. Lastly, CaV channels can be therapeutically inhibited to manage pain, epilepsy, arrythmia, high blood pressure, and alternatively, activated to treat heart failure. Surprisingly, both of these effects (channel activation and inactivation) can be elicited by medicines binding a common extracellular binding site on the channel. Conversely, unintended blockade of cardiac hERG potassium channels by otherwise useful therapeutics cause 90% of drug induced long-QT syndrome, a potentially lethal cardiac arrhythmia. All of these chemical binding events rely on aromatic rich binding sites formed by the side-chains of phenylalanine and tyrosine residues in CaV and hERG channels. To better understand these chemical interactions, we have developed a high-resolution method that allows for energetic and nuanced dissection of these aromatics within the CaV and KV drug binding aromatic boxes in the environment of mammalian cells. The successful execution of this research program will provide cutting edge training opportunities, advance the molecular understanding of channel gating, and will reveal the binding modes of clinical drugs with high therapeutic value.

电压门控离子通道在神经和肌肉的可激发细胞中塑造电信号传导。钠（NAV） 和钙通道（CAV）驱动膜去极化并通过门激活第二通道通路 其同名离子的细胞进入。在骨骼和心脏细胞中，CAV通道会触发肌肉收缩。 电压门控钾通道（KV）允许钾离子从细胞内释放到驱动 膜复极化。在一致的情况下，这些渠道为思想，感知， 和收缩。人类电压门控通道的高分辨率蛋白质结构现在提供第一个 瞥见它们在细胞环境中可能采用的姿势类型。但是，了解最终 这些蛋白质的外观与它们如何支持生理机制之间的联系是一个主要挑战 将需要创新的方法。首先，在结构实验中不存在跨膜电压 在本质上非生理环境中描述电压门控通道。因此，我们正在发展 光化学的“订书”方法是共价捕获活细胞膜中的高价值蛋白构象 在纯化结构确定之前。此外，我们已经开始确定渠道的机制 通过在形成的跨膜段中的肽主链处引入改良化学作用。 电压传感器和通道门。在细胞设置中，离子通道也是转导的关键放大器 途径。在战斗或战斗反应中，例如，Cav1.2的近瞬时磷酸化 通道会导致更快，持续的通道开口，从而导致更强大和快速的心率。然而 该过程的绝对速度和复杂性是实验解析个体分子的挑战 导致通道门控修改的事件。我们描述了启用光的新验证的方法 受控的，特定于现场的磷酸化，以仔细解构和识别关键步骤和参与者 是这个过程。最后，可以在治疗上抑制CAV通道以控制疼痛，癫痫，雅利氏病，高 血压，或者被激活以治疗心力衰竭。令人惊讶的是，这两种效果（渠道 激活和灭活）可以通过结合共同细胞外结合位点的药物引起 渠道。相反，通过有用的意外封锁心脏HERG钾通道 治疗药引起90％的药物诱导长QT综合征，这是一种潜在的致命心律不齐。所有这些 化学结合事件依赖于由苯丙氨酸和 CAV和HERG频道中的酪氨酸残留物。为了更好地了解这些化学相互作用，我们有 开发了一种高分辨率方法，该方法允许这些芳香剂的能量和细微的解剖 哺乳动物细胞环境中的CAV和KV药物结合芳族盒。成功的执行 该研究计划将提供尖端培训机会，提高分子理解 通道门控的，并将揭示具有高治疗价值的临床药物的结合模式。