BRAIN CALCIUM AND CALMODULIN-DEPENDENT PROTEIN KINASE

脑钙和钙调蛋白依赖性蛋白激酶

• 批准号: 3397710
• 负责人: MARY B KENNEDY
• 金额: $ 23.8万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-07-01 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3397710
• 关键词: allosteric site; antibody specificity; autoradiography; brain metabolism; calcium flux; calmodulin dependent protein kinase; chemical structure function; cytoskeleton; electron microscopy; enzyme structure; gel electrophoresis; genetic manipulation; high performance liquid chromatography; hippocampus; histochemistry /cytochemistry; immunochemistry; immunological substance; laboratory mouse; laboratory rabbit; laboratory rat; microtubules; monoclonal antibody; neurotransmitters; phosphorylation; radioimmunoassay; recombinant DNA; scintillation spectrometry; synapses; synaptic vesicles

We are studying the molecular structure and function of synaptic connections between neurons in the central nervous system. Derangements in the regulation of these connections are an important part of the pathology of several neurological and mental diseases including epilepsy, Alzheimer's disease, schizophrenia, and depression. Many neurotransmitters and neurohormones regulate synaptic function by altering intracellular levels of calcium ion. We are studying the mechanisms by which these fluctuations in calcium levels alter synaptic function. We will focus on the study of a synaptic regulatory pathway that has as its central element an abundant, brain-specific calcium and calmodulin-dependent protein kinase. This kinase is a large oligomer of two distinct but homologous catalytic subunits called alpha and beta. In the forebrain, including the hippocampus, cortex and striatum, the kinase is extremely abundant (1% of total protein) and is composed mainly of alpha subunits. It is a major component of synapses and is concentrated in a cytoskeletal structure called the postsynaptic density. When activated by a brief rise in calcium concentration, the kinase phosphorylates itself and then remains active to phosphorylate other proteins even after the calcium concentration falls. We will test the hypothesis that this is a mechanism by which long-lasting changes in synaptic function are generated following brief bursts of synaptic activity. We will determine the structure of the autophosphorylation sites by recombinant DNA and biochemical methods, then characterize the brain phosphatases responsible for dephosphorylation of each of these sites. We will raise antibodies that specifically recognize the autophosphorylated sites on the kinase and others that recognize the phosphorylated form of kinase substrates. We will use these to study, with high spatial and temporal resolution, the physiological circumstances under which the kinase is activated and specific substrates become phosphorylated. We will continue a study of the association of the kinase with the cytoskeleton by biochemical and recombinant DNA techniques. Our goal in the next few years is to clarify the possible regulatory functions of this calcium-dependent protein kinase system. Our long-term goal is to correlate information about this pathway with similar information about other calcium regulated pathways in order to understand the concerted responses to changing calcium levels in CNS neurons.

我们正在研究突触的分子结构和功能 中枢神经系统中神经元之间的连接。 这些连接在监管上的错乱是一种 是几种神经性和非神经性疾病的重要病理部分 精神疾病，包括癫痫，阿尔茨海默氏症， 精神分裂症和抑郁症。许多神经递质和 神经激素通过改变细胞内信号调节突触功能 钙离子水平。我们正在研究通过什么机制 钙水平的这些波动会改变突触功能。我们 将专注于对突触调节途径的研究 作为它的中心元素，丰富的，大脑特有的钙和 钙调蛋白依赖的蛋白激酶。这种激酶是一种很大的 两个不同但同源的催化亚基的齐聚物，称为 阿尔法和贝塔。在前脑，包括海马体， 大脑皮层和纹状体的肌动蛋白含量极其丰富(1% 总蛋白)，主要由α亚基组成。这是一个 是突触的主要组成部分，集中在 细胞骨架结构称为突触后密度。什么时候 在短暂的钙浓度上升的激活下，该激酶 将自身磷酸化，然后保持活性以进行磷酸化 其他蛋白质即使在钙浓度下降后也是如此。我们会 检验这一假设，即这是一种长期持续的 突触功能的变化是在短暂的突触后产生的 突触活动。我们将确定 重组DNA和生化研究中的自动磷酸化位点 方法，然后表征负责脑内磷酸酶 这些位点中的每一个都去磷酸化。我们会提高抗体 它特别识别细胞上的自动磷酸化位点 激酶和其他识别磷酸化形式的 酶底物。我们将利用这些进行研究，具有较高的空间性 和时间分辨率，下面的生理环境 激活剂被激活，特定的底物变成 磷酸化。我们将继续研究两国之间的联系。 利用生物化学和重组技术将细胞骨架与激活剂结合 DNA技术。我们未来几年的目标是澄清 这种钙依赖蛋白可能的调节功能 激酶系统。我们的长期目标是将信息关联起来 有关这一途径的信息以及关于其他钙离子的类似信息 调控通路，以了解协同 中枢神经系统神经元对钙水平变化的反应。