STRUCTURAL THERMODYNAMICS OF HUMAN APOLIPOPROTEIN C 1

人载脂蛋白 C 1 的结构热力学

• 批准号: 6330179
• 负责人: Olga Gursky
• 金额: $ 11.38万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-12-15 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6330179
• 关键词: X ray crystallography; amphiphilicity; apolipoproteins; calorimetry; chemical models; chemical stability; circular dichroism; conformation; crystallization; protein folding; protein sequence; protein structure function; site directed mutagenesis; structural biology; synthetic protein; thermodynamics

The long-term objective of this project is to determine in molecular detain the energetic-structure-function relationship in exchangeable apolipoproteins, thereby providing an insight into molecular mechanisms of their action in the pathogenesis of atherosclerosis and other apolipoprotein-related disorders. Exchangeable apolipoproteins are water- soluble protein components of lipoproteins that mediate lipid and cholesterol transport and metabolism and play crucial roles in the pathogenesis of atherosclerosis, coronary heart disease, stroke, and other major human disorders including several forms of systemic and cerebral amyloidosis. Structural adaptability of apolipoproteins to heterogenous lipoprotein complexes and to plasma is absolutely essential for their functions, and has to be understood in detain in order to elucidate molecular mechanisms of apolipoprotein action in normal and in diseased states. The proposed work addresses this long-term goal through studies of the energetics, structure and folding pathway of the smallest human plasma apolipoprotein C-1 (apoC-1, 6 kDa). The ability of apoC-1 to activate lecitin: cholesterol acyltransferase (LCAT) may account for normal plasma levels of cholesterol esters in subjects with deficiency of the major LCAT activator, apoA-1. ApoC-1 delays the clearance of potentially atherogenic triglyceride-rich particles by inhibiting their uptake via the apoE-mediated low-density low-density lipoprotein receptor- related pathway. Thermodynamic and structural analyses of synthetic human apoC-1 and a series of its site-specific mutants targeted towards the predicted amphipathic alpha-helical regions will be carried out by using a combination of far-UV circular dichroism spectroscopy, differential scanning calorimetry, and x-ray diffraction methods. Such analysis will 1) dissect the folding pathway of lipid-free apoC-1 in solution, from partly folded monomeric to fully folded oligomeric or lipid-bound state; 2) determine, at the level of individual amino acids, the critical factors for the stability and cooperatively of the amphipathic alpha-helical structure in apoC-1; 3) determine the conformation apoC-1 in various self- associated status, such as 2D filaments and 3D crystals, to provide models fir a variety of functional apolipoprotein conformations. The results of this analysis will provide the energetic and structural basis for understanding mechanisms of functional apolipoprotein reactions and will help to understand their amyloidogenic properties, thereby leading to identification of rational therapeutic targets in a variety of apolipoprotein-related disorders.

该项目的长期目标是确定分子 保留可交换的能量-结构-功能关系 载脂蛋白，从而深入了解载脂蛋白的分子机制 它们在动脉粥样硬化和其他疾病发病机制中的作用 载脂蛋白相关疾病。可交换性载脂蛋白是水 脂蛋白的可溶性蛋白质成分，介导脂质和 胆固醇的转运和代谢在胆固醇的运输和代谢中发挥着至关重要的作用 动脉粥样硬化、冠心病、中风等疾病的发病机制 主要人类疾病，包括多种形式的全身性和脑部疾病 淀粉样变性。载脂蛋白对异源的结构适应性 脂蛋白复合物和血浆对于它们来说是绝对必要的 功能，并且必须在拘留中理解才能阐明 正常和患病的载脂蛋白作用的分子机制 州。拟议的工作通过研究解决了这一长期目标 最小人类的能量学、结构和折叠途径 血浆载脂蛋白 C-1（apoC-1，6 kDa）。 apoC-1 的能力 激活卵磷脂：胆固醇酰基转移酶 (LCAT) 可能是 缺乏胆固醇的受试者血浆胆固醇酯水平正常 主要的 LCAT 激活剂 apoA-1。 ApoC-1 延迟清除 潜在致动脉粥样硬化的富含甘油三酯的颗粒通过抑制其 通过 apoE 介导的低密度低密度脂蛋白受体摄取 相关途径。人造人的热力学和结构分析 apoC-1 及其一系列针对 预测的两亲性 α 螺旋区域将通过使用进行 远紫外圆二色光谱、微分光谱的组合 扫描量热法和X射线衍射法。这样的分析将 1) 剖析溶液中无脂apoC-1的折叠途径，部分 折叠单体至完全折叠寡聚或脂质结合状态； 2） 在单个氨基酸水平上确定关键因素 为了两亲性α螺旋的稳定性和协同性 apoC-1 的结构； 3) 确定apoC-1在各种自体中的构象 相关状态，例如 2D 灯丝和 3D 晶体，以提供模型 fir有多种功能性载脂蛋白构象。结果 这一分析将为 了解功能性载脂蛋白反应的机制和意愿 有助于了解它们的淀粉样蛋白生成特性，从而导致 确定各种合理的治疗靶点 载脂蛋白相关疾病。