Natural Agonists of Ryanodine Receptors: Structure-function Relationship and Antiarrhythmic Properties

兰尼定受体的天然激动剂：结构-功能关系和抗心律失常特性

• 批准号: 9650244
• 负责人: Hector H Valdivia
• 金额: $ 46.18万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9650244
• 关键词: Active Sites; Acute; Adrenergic alpha-Antagonists; Affinity; Agonist; Amino Acids; Animal Model; Animals; Arrhythmia; Binding; Binding Sites; Calcium; Calcium Channel; Cardiac Myocytes; Cardiomyopathies; Catecholaminergic Polymorphic Ventricular Tachycardia; Cell physiology; Cells; Cellular Membrane; Central Core Myopathy; Characteristics; Charge; Communities; Complex; Disadvantaged; Dissociation; Dose; Enzyme Activation; Functional disorder; Generations; Goals; Heart; Heart failure; Hyperactive behavior; Ions; Kinetics; Ligands; Location; Malignant hyperpyrexia due to anesthesia; Maps; Metabolic; Molecular; Muscle Cells; Myocardium; Names; Penetration; Peptides; Pharmacology; Plant alkaloid; Play; Property; Proteins; Receptor Activation; Recombinants; Regulation; Research; Role; RyR2; Ryanodine; Ryanodine Receptor Calcium Release Channel; Ryanodine Receptors; Sarcoplasmic Reticulum; Scorpion Venoms; Scorpions; Signal Transduction; Skeletal Muscle; Specificity; Speed; Structure-Activity Relationship; Syndrome; Testing; Transcriptional Regulation; Venoms; Ventricular; dipole moment; experimental study; functional group; imperatoxin A; multidisciplinary; mutant; novel; prevent; programs; receptor; receptor function; receptor structure function; success; thermostability

ABSTRACT Ryanodine receptors (RyR) are sarcoplasmic reticulum Ca2+ release channels that play a critical role in Ca2+ signaling of excitable and non-excitable cells. RyRs owe their name to the fact that they were characterized in great part thanks to ryanodine, a plant alkaloid that binds to RyRs with high affinity and specificity. Ryanodine has been an invaluable ligand of RyRs, but its functional effects are complex and hamper its use in cellular studies. In search of novel ligands that could overcome some of the functional and structural disadvantages of ryanodine, we found in the venom of selected scorpions a set of peptide ligands, termed calcins, displaying high affinity and exquisite selectivity against RyRs. The defining characteristic of calcins is their capacity to stabilize RyR openings in a long-lasting subconducting state. This effect is nearly analogous to that of ryanodine, but unlike ryanodine, calcins bind rapidly to RyRs (fast association rate), freely dissociate from their binding site (reversible effect), display a dose- and sequence-variable effect, and are amenable for derivatization without undergoing major loss in receptor affinity. Calcins also modulate intracellular Ca2+ in intact cardiomyocytes with remarkable speed and with several degrees of potency, thus entering the field as the first cell-penetrating peptides (CPP) RyR-specific Ca2+ mobilizer of high dynamic range. This research program will characterize first and then exploit this novel group of peptide ligands to unravel fundamental mechanisms of RyR function at the molecular, cellular and whole heart level. Our multidisciplinary program, with well-defined deliverables and milestones, may be enveloped in two specific aims. In the first aim, we will first identify and modify the structural domains of calcins involved in RyR recognition and cell penetration to generate a group of functionally diverse CPPs capable of modulating RyR function with wide dynamic range and of delivering cargo to the interior of cardiomyocytes. In the second aim, we will use native and mutant calcins on intact cardiomyocytes, Langendorff-perfused working hearts and intact animals to create acute or sustained periods of RyR hyperactivity and reveal mechanisms of RyR gating, SR Ca2+ load and Ca2+-triggered arrhythmias. These studies use animal models of catecholaminergic polymorphic ventricular tachycardia to develop a novel paradigm for the treatment of calcium-dependent arrhythmias; results may be applied to other cardiomyopathies where controlled unloading of SR Ca2+ may be desirable.

摘要 Ryanodine受体（RyR）是肌浆网Ca 2+释放通道，在肌浆网钙通道的形成和维持中起关键作用。 可兴奋和非兴奋细胞的Ca 2+信号传导。RyR的名字来源于这样一个事实，即它们的特征是 这主要归功于ryanodine，一种以高亲和力和特异性结合RyR的植物生物碱。Ryanodine有 作为RyRs的一种非常宝贵的配体，但其功能作用很复杂，阻碍了其在细胞研究中的应用。在 寻找新的配体，可以克服一些功能和结构的缺点，兰尼碱，我们 在选定的蝎子毒液中发现了一组肽配体，称为钙蛋白，显示出高亲和力， 对RyR的精确选择性。钙蛋白的定义特征是它们稳定细胞中RyR开口的能力。 一种持久的亚导电状态。这种作用几乎类似于ryanodine，但与ryanodine不同的是， 与RyR快速结合（快速结合速率），从其结合位点自由解离（可逆效应），显示剂量- 和序列可变效应，并且适合于衍生化而不经历受体亲和力的重大损失。 钙蛋白还以显著的速度调节完整心肌细胞中的细胞内Ca 2+，并具有几种程度的抑制作用。 效力，从而进入该领域的第一个细胞穿透肽（CPP）RyR特异性钙动员剂的高 动态范围这项研究计划将首先表征，然后利用这组新的肽配体， 在分子、细胞和整个心脏水平上阐明RyR功能的基本机制。我们的多学科 一个具有明确的可交付成果和里程碑的计划可能包含两个具体目标。第一个目标，我们 将首先识别和修饰参与RyR识别和细胞渗透的钙蛋白的结构域， 产生一组功能多样的CPP，其能够以宽的动态范围调节RyR功能， 将货物运送到心肌细胞内部。在第二个目标中，我们将使用天然和突变的钙蛋白对完整的 心肌细胞，Langendorff灌注的工作心脏和完整的动物，以产生急性或持续的RyR 并揭示了RyR门控、SR Ca ~（2+）负荷和Ca ~（2+）触发心律失常机制。这些研究采用了 儿茶酚胺能多形性室性心动过速的动物模型，以开发新的治疗范例 钙依赖性心律失常;结果可能适用于其他心肌病， SR Ca 2+可能是理想的。