Regulation of Calcium Channels by Calmodulin

钙调蛋白对钙通道的调节

• 批准号: 7613432
• 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/7613432
• 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; 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; Structure; Tail; Testing; Therapeutic; Timothy syndrome; Ursidae Family; base; human disease; luminescence resonance energy transfer; prevent; programs; research study; skeletal; 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，以调节每种肌肉的功能。该法规的组成部分是通过通道本身进入钙进入这些通道的反馈抑制和反馈激活这些通道。这些反馈机制，依赖钙的失活（CDI）和钙依赖性促进（CDF），都取决于钙结合与钙调蛋白的结合以及构象的变化，导致通道内钙调蛋白结合位点的变化。仍然未知如何通过相同的蛋白质在通道C末端的复杂结合位点调节这两种机制。该提议的假设是，反馈调节是通过转移Ca2+频道α-硫酸盐C末端结构域内钙调蛋白的构象和结合​​而发生的。我们将通过三个特定目的来检验这一假设：AIM 1将使用高精度，基于灯笼的能量传递距离测量钙调蛋白之间的钙质形式中的钙调蛋白在其APO形式中的构象，而没有钙占用率。裂片将通过距离测量到通道上的特定识别序列进行定位。此外，X射线晶体学研究将确定与识别序列复合的Apo-钙统蛋白的原子分辨率结构。 AIM 2将利用类似的方法来确定每个钙调蛋白叶的结合位点及其在CDI期间发生的部分和完整的Ca2+占用过程中的构象。 AIM 3将使用类似的方法在存在已知会影响CDI和CDF的突变的情况下确定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期间钙调蛋白的构象和位置的确切变化，并对如何启动这些调节功能进行批判性理解。该提案中研究的蛋白质调节心跳和骨骼肌收缩。这些渠道在人类疾病中至关重要。在心脏中，它们是钙阻滞剂的靶标，通常用于预防心脏病发作。拟议的研究提供了有关该通道如何调节的信息，因此，为诸如长QT综合征，高钾血症周期性瘫痪，恶性高温和蒂莫西综合症等疾病提供更好疗法的机会。