Energetics-Structure-Function Relationship in Lipoproteins

我们的长期目标是确定载脂蛋白的能量-结构-功能关系， 脂蛋白，以更好地了解脂蛋白作用的分子机制，在正常和 动脉粥样硬化状态重点是载脂蛋白在溶液中的折叠、结构和稳定性 和脂蛋白，特别是模型盘状HDL和人血浆或重构LDL。的 血浆中HDL、LDL及其亚类之间的平衡决定了发生 心血管疾病和中风。脂蛋白的结构稳定性对其功能至关重要，然而， 其稳定的精确机制是未知的。此外，HDL和LDL被广泛地重新建模 例如，LDL在动脉壁中的融合是代谢早期的关键事件。 动脉粥样硬化然而，HDL和LDL重构和融合的分子机制还不清楚。 我们的研究旨在详细了解其能量和结构基础 HDL和LDL的稳定性和重塑。我们对重组盘状HDL的初步研究表明， 揭示了一种新的脂蛋白稳定的动力学机制;他们表明，脂蛋白不稳定 和蛋白质解离导致涉及高能量势垒的粒子融合。我们证明， 类似的基于融合的动力学机制赋予血浆HDL和LDL稳定性，从而提供了一种 脂蛋白稳定的普遍天然策略。在拟议的工作中，我们将获得详细的分子 盘状HDL动力学稳定性的决定因素。重点将是蛋白质大小的作用， 疏水性，电荷残基分布，和脂质组成的磁盘的稳定性，重点是 基于apoA-1的蛋白质和肽。将复溶受控组成的盘状HDL， 通过使用集成的光谱、量热和电子显微镜方法进行分析。动力学 蛋白质-脂质缔合的，这将通过吸收和圆二色光谱分析， 有助于确定蛋白质一级和二级结构对脂质结合途径的作用。 将获得LDL稳定性和融合的分子决定因素，重点是 蛋白质和脂质氧化，脂质组成，和载脂蛋白B C-末端截短的结构和稳定性， 血浆或重组脂蛋白。HDL和LDL的能量和结构分析，例如 这项建议可能导致鉴定促进或抑制促动脉粥样硬化脂蛋白的化合物 例如融合等转化，从而有助于开发冠状动脉疾病的新疗法， 其他脂蛋白相关疾病。载脂蛋白和抗融合作用的比较研究 肽还可以帮助设计具有优化的抗病毒特性的基于载脂蛋白的肽， 确定LDL稳定性的关键决定因素可能有助于设计基于LDL的药物载体。

Our long-term goal is to determine the energetic-structure-function relationship in apolipoproteins and lipoproteins to gain a better insight into the molecular mechanisms of lipoprotein action in normal and in atherosclerotic states. The emphasis is on the folding, structure and stability of apolipoproteins in solution and on lipoproteins, particularly on model discoidal HDL and human plasma or reconstituted LDL. The balance between HDL, LDL and their subclasses in plasma determines the probability of developing cardiovascular disease and stroke. Structural stability of lipoproteins is essential for their functions, yet the precise mechanism of their stabilization is unknown. Furthermore, HDL and LDL are extensively re-modeled by plasma factors during metabolism; for example, LDL fusion in the arterial wall is a key event in early atherosclerosis. However, the molecular mechanisms of HDL and LDL remodeling and fusion are not wellunderstood. Our research is aimed at detailed understanding of the energetic and structural basis underlying stability and re-modeling of HDL and LDL. Our preliminary studies of reconstituted discoidal HDL have revealed a novel kinetic mechanism of lipoprotein stabilization; they showed that lipoprotein destabilization and protein dissociation lead to particle fusion that involves high energy barriers. We demonstrated that a similar fusion-based kinetic mechanism confers stability to plasma HDL and LDL, and thus provides a universal natural strategy for lipoprotein stabilization. In the proposed work we will obtain detailed molecular determinants for the kinetic stability of discoidal HDL. The focus will be on the role of protein size, hydrophobicity, charge residue distribution, and lipid composition in the disks stability, with an emphasis on apoA-1-based proteins and peptides. Discoidal HDL of controlled composition will be reconstituted and analyzed by using an integrated spectroscopic, calorimetric, and electron microscopic approach. The kinetics of protein-lipid association, which will be analyzed by absorption and circular dichroism spectroscopy, will help to determine the role of the protein primary and secondary structure on the lipid binding pathway. Molecular determinants for LDL stability and fusion will be obtained, with an emphasis on the effects of protein and lipid oxidation, lipid composition, and apoB C-terminal truncations on the structure and stability of plasma or reconstituted lipoproteins. The energetic and structural analysis of HDL and LDL, such as this proposal, may lead to identification of compounds that promote or inhibit pro-atherogenic lipoprotein transformations such as fusion, and thereby help to develop new therapies for coronary artery disease and other lipoprotein-related disorders. Comparative studies of antifusogenic effects of apolipoproteins and peptides may also help to design apolipoprotein-based peptides with optimized antiviral properties, while identification of key determinants for LDL stability may help to design LDL-based drug carriers.