Regulation of Calcium Channels by Calmodulin

钙调蛋白对钙通道的调节

• 批准号: 7185205
• 负责人: STEEN E PEDERSEN
• 金额: $ 38.38万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2011-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7185205
• 关键词: Affect; Binding; Binding Sites; C-terminal; Calcium; Calcium Binding; Calcium Channel; Calmodulin; Cardiac; Complex; Crystallography; Cysteine; Detection; Disease; EF Hand Motifs; Energy Transfer; Engineering; Feedback; Goals; Heart; Hyperkalemic periodic paralysis; Hypokalemic periodic paralysis; Label; Lanthanoid Series Elements; Lobe; Localized; Location; Long QT Syndrome; Malignant hyperpyrexia due to anesthesia; Measurement; Measures; Mediating; Modeling; Molecular; Molecular Conformation; Muscle; Muscle Contraction; Mutation; Myocardial Infarction; N-terminal; Peptide Fragments; Process; Proteins; Regulation; Relative (related person); Research; Research Personnel; Resolution; Rest; Roentgen Rays; Site; Skeletal Muscle; Skeletal system; Structure; Tail; Testing; Therapeutic; Timothy syndrome; Ursidae Family; base; human disease; luminescence resonance energy transfer; prevent; programs; research study; tool; voltage

DESCRIPTION (provided by applicant): Voltage-gated calcium channels of the heart and of skeletal muscle are highly regulated to tune the function of each type of muscle. An integral part of the regulation is feedback inhibition and feedback activation of these channels by calcium entry through the channel itself. These feedback mechanisms, calcium-dependent inactivation (CDI) and calcium-dependent facilitation (CDF), both depend on calcium binding to calmodulin and a change in conformation leading to a shift in the calmodulin binding site within the channel. It is still unknown how these two mechanisms can be regulated through the same protein at a complex binding site of the C-terminal tail of the channel. The hypothesis of this proposal is that feedback regulation occurs by shifting the conformation and binding of calmodulin within the C-terminal domain of the Ca2+-channel alpha-subunit. We will test this hypothesis through three specific aims: Aim 1 will examine the conformation of calmodulin in its apo-form, with no calcium occupancy, using high-precision, lanthanide-based energy transfer distance measurements between the lobes of calmodulin. The lobes will be localized by distance measurements to specific recognition sequences on the channel. In addition, X-ray crystallographic studies will determine the atomic-resolution structure of apo-calmodulin complexed with the recognition sequences. Aim 2 will utilize similar approaches to determine the binding sites for each calmodulin lobe and its conformation during partial and complete Ca2+ occupancy that occur during CDI. Aim 3 will determine conformation and sites of calmodulin binding in the presence of mutations known to affect CDI and CDF, using similar approaches. The results of these experiments will reveal the exact changes in conformation and location of calmodulin that occur during CDI and CDF and to provide a critical understanding of how these regulatory functions are initiated. The proteins under study in this proposal regulate the heart beat and skeletal muscle contraction. These channels are critically important in human disease; in the heart they are the targets of calcium blockers, which are routinely used to prevent heart attack. The proposed research provides information on how this channel is regulated, and, therefore, an opportunity to provide better therapeutics for diseases such as long QT syndrome, hyperkalemic periodic paralysis, malignant hyperthermia, and Timothy's syndrome.

描述（由申请人提供）：心脏和骨骼肌的电压门控钙通道受到高度调节，以调节每种类型肌肉的功能。调节的一个组成部分是通过钙通过通道本身进入这些通道的反馈抑制和反馈激活。这些反馈机制，钙依赖性失活（CDI）和钙依赖性促进（CDF），都依赖于钙与钙调蛋白的结合以及导致通道内钙调蛋白结合位点发生变化的构象变化。目前尚不清楚这两种机制如何通过通道 C 末端尾部的复杂结合位点上的同一蛋白质进行调节。该提议的假设是，反馈调节是通过改变 Ca2+ 通道 α 亚基 C 端结构域内钙调蛋白的构象和结合​​来发生的。我们将通过三个具体目标来检验这一假设：目标 1 将使用高精度、基于镧系元素的钙调蛋白叶之间的能量转移距离测量来检查钙调蛋白的 apo 形式的构象，没有钙占据。波瓣将通过距离测量来定位到通道上的特定识别序列。此外，X 射线晶体学研究将确定与识别序列复合的脱辅基钙调蛋白的原子分辨率结构。目标 2 将利用类似的方法来确定 CDI 期间发生的部分和完全 Ca2+ 占据期间每个钙调蛋白叶的结合位点及其构象。目标 3 将使用类似的方法确定已知影响 CDI 和 CDF 的突变存在时钙调蛋白结合的构象和位点。这些实验的结果将揭示 CDI 和 CDF 期间发生的钙调蛋白构象和位置的确切变化，并提供对这些调节功能如何启动的批判性理解。该提案中研究的蛋白质调节心跳和骨骼肌收缩。这些通道对于人类疾病至关重要。在心脏中，它们是钙阻滞剂的目标，钙阻滞剂通常用于预防心脏病发作。拟议的研究提供了有关如何调节该通道的信息，因此有机会为长 QT 综合征、高钾性周期性麻痹、恶性高热和蒂莫西综合征等疾病提供更好的治疗方法。