Mechanisms of Ion Channel Activity

离子通道活性机制

• 批准号: 6798223
• 负责人: KARL L MAGLEBY
• 金额: $ 36.74万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-09-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6798223
• 关键词: Xenopus oocyte; calcium flux; cell line; cell membrane; chemical binding; chemical kinetics; intermolecular interaction; ion transport; membrane potentials; potassium channel; potassium ion; protein structure function; voltage /patch clamp; voltage gated channel

DESCRIPTION (provided by applicant):Ion channels are proteins that control the passive flux of ions through cell membranes by opening and closing (gating) their pores. This proposal seeks to continue work on investigating the fundamental mechanisms by which ion channels gate. The current focus is the large conductance Ca 2+-and voltage activated K+ (BK) channel, which plays a key role in many physiological processes, including control of muscle contraction, regulation of synaptic transmission, and integration of information in neurons. Although much progress has been made towards understanding how Ca 2+ and voltage activate BK channels, and how accessory beta1subunits modulate this activity, many basic questions remain. To work towards answering these questions, cloned BK channels will be expressed in Xenopus oocytes and HEK293 cells, currents will be recorded from single channels with the patch clamp technique, and the data will be analyzed with advanced techniques to determine gating mechanism. The initial hypothesis to be tested is that multiple Ca 2+- and voltage-dependent regulatory mechanisms act jointly to control the opening-closing transitions (gating) of BK channels. To test this hypothesis, the first specific aim will: 1) determine the contributions, including any cooperative interactions, of each of the five proposed Ca 2+-- dependent regulatory mechanisms to the gating of BK channels. Cooperatively it will be resolved by studying the Ca2+-dependent regulatory mechanisms in isolation and in various combinations. The second specific aim will: 2) develop a comprehensive kinetic gating mechanism for BK channels that incorporates all of the known Ca 2+and voltage-dependent regulatory mechanisms and their cooperative interactions. The gating mechanism will be formulated in terms of a large multi-state multi-tiered model that specifies the states, the transitions among the states, the rate constants for the transitions, modulation of the rate constants by Ca 2+ and voltage, and any cooperative interactions involved in the gating. The third specific aim will: 3) determine the mechanism by which beta1subunits modulate the gating of BK channels. This will be done by identifying the particular steps in the kinetic gating mechanism that are modified by beta1subunits. Understanding how ion channels gate their pores, the goal of this research, will facilitate the comprehension, diagnosis, and treatment of diseases associated with defective ion channels (channelopathies).

描述（由申请人提供）：离子通道是通过打开和关闭（门控）其孔来控制离子通过细胞膜的被动通量的蛋白质。该提案旨在继续研究离子通道门控的基本机制。目前的焦点是大电导Ca 2+- 和电压激活的K+ (BK) 通道，它在许多生理过程中发挥着关键作用，包括肌肉收缩的控制、突触传递的调节以及神经元中信息的整合。尽管在理解 Ca 2+ 和电压如何激活 BK 通道以及辅助 beta1 亚基如何调节这种活动方面已经取得了很大进展，但仍然存在许多基本问题。为了回答这些问题，克隆的 BK 通道将在爪蟾卵母细胞和 HEK293 细胞中表达，使用膜片钳技术从单个通道记录电流，并使用先进技术分析数据以确定门控机制。要测试的最初假设是多种 Ca 2+- 和电压依赖性调节机制共同作用来控制 BK 通道的打开-关闭转换（门控）。为了检验这一假设，第一个具体目标将：1) 确定所提出的五种 Ca 2+ 依赖性调节机制中每一种对 BK 通道门控的贡献，包括任何合作相互作用。通过单独和各种组合研究 Ca2+ 依赖性调节机制，可以合作解决该问题。第二个具体目标是：2) 开发一种全面的 BK 通道动力学门控机制，其中包含所有已知的 Ca 2+ 和电压依赖性调节机制及其协作相互作用。门控机制将根据大型多状态多层模型来制定，该模型指定状态、状态之间的转换、转换的速率常数、Ca 2+ 和电压对速率常数的调制以及任何门控中涉及的合作相互作用。第三个具体目标是：3) 确定 beta1 亚基调节 BK 通道门控的机制。这将通过识别由 beta1 亚基修饰的动力学门控机制中的特定步骤来完成。这项研究的目标是了解离子通道如何控制其毛​​孔，这将有助于理解、诊断和治疗与离子通道缺陷（通道病）相关的疾病。