Impact of self-association on structure and function of apolipoprotein A-I

描述(由申请人提供)：这项资助提案的主要目标是证明载脂蛋白A-I(apoA-I)的自结合是蛋白质的一个功能特征，并且其天然自结合状态的改变涉及导致疾病的机制。自结合是几种可交换载脂蛋白的无脂形式的固有属性，包括载脂蛋白A-I，高密度脂蛋白的主要蛋白质成分，以及公认的抗动脉粥样硬化因子。单体无脂apoA-I被认为是具有生物活性的物种。但异常情况，如特定的突变或氧化，会产生一种改变的自我关联状态，这可能导致载脂蛋白A-I功能障碍。虽然自结合在一些载脂蛋白中的功能作用已经确定，但由于本项目解决和克服的技术限制，以前还没有研究过自结合对载脂蛋白A-I功能的影响。ApoA-I的TRPs被Phes(？W-apoA-I)取代，导致了异常大和稳定的自伴生物种。至少可以分离出四个自相关的W-apoA-I物种，并将在这里作为自关联的模型来分析它在确定apoA-I的结构、功能和对导致功能障碍的机制的敏感性方面所起的作用。本项目要研究的三个重叠领域是：1.确定自关联在apoA-I结构的所有水平上的影响，从二级到第四级。将确定apoA-I自结合所涉及的分子间相互作用的性质，并将确定导致自结合程度增加的脂质结合效率丧失的结构细节。这种结构知识将有助于理解蛋白质自结合状态改变影响其生物学功能的机制。2.研究在高密度脂蛋白的生物发生过程中，无脂apoA-I的自结合状态如何影响其作为细胞释放的脂类受体的功能。不同的apoA-I自结合物种激活不同细胞膜转运体介导的脂质释放的效率将被确定。包括ABCA1，这是apoA-I抗动脉粥样硬化作用的主要机制。A.证明无脂apoA-I的自结合状态调节蛋白质对导致功能障碍的机制的敏感性。将测试不同的自我相关物种对与动脉粥样硬化和糖尿病的发病机制有关的反应的脆弱性。自我结合在保护载脂蛋白A-I不受促进淀粉样原纤维形成的条件的影响方面的可能作用也将被评估，淀粉样纤维形成是动脉粥样硬化进展的一个促成机制。这些研究对公众健康具有非常重要的意义，因为确定载脂蛋白A-I的一个新的功能方面，这是已知的最重要的抗动脉粥样硬化因子之一，具有制定新的治疗方法和开发新的生物标记物评估心血管疾病风险的潜力。

DESCRIPTION (provided by applicant): The primary goal of this grant proposal is to demonstrate that self-association of apolipoprotein A-I (apoA-I) is a functional feature of the protein and that alteration of its native state of self-association is involved in mechanisms leading to disease. Self-association is an inherent property of the lipid-free forms of several exchangeable apolipoproteins, including apoA-I, the main protein component of HDL and an established antiatherogenic factor. Monomeric lipid-free apoA-I is believed to be the biologically active species. But abnormal conditions, such as specific mutations or oxidation, produce an altered state of self-association that may contribute to apoA-I dysfunction. Although the functional role of self-association in some apolipoproteins has been established, the influence of self-association on apoA-I function has not been studied before because of technical limitations that are addressed and overcome in this project. Replacement of apoA-I's Trps with Phes (¿W-apoA-I) leads to unusually large and stable self-associated species. At least four self-associated species of ¿W-apoA-I can be isolated and will be used here as a model of self-association to analyze its role in determining apoA-I's structure, function, and susceptibility to mechanisms leading to dysfunction. The three overlapping areas to be investigated in this project are: 1. Define the effects of self-association at all levels of apoA-I structure, from secondary to quaternary. The nature of the inter-molecular interactions that are involved in apoA-I self-association will be established and the structural details underlying the loss of lipid-binding efficiency for increasing degrees of self- association will be determined. This structural knowledge will help to understand the mechanisms whereby alteration of the protein self-association state affects its biological function. 2. Characterize how the self-association state o lipid-free apoA-I affects its function as recipient of lipids released from cells in the biogenesisof HDL. The efficiency of different apoA-I self-associated species in activating lipid release mediated by different cell membrane transporters will be determined. Including ABCA1, which is the primary mechanism underlying the anti-atherogenic function of apoA-I. a. Demonstrate that the self-association state of lipid-free apoA-I modulate the protein susceptibility to mechanisms leading to dysfunction. The vulnerability of different self-associated species to reactions that are implicated in the pathogenesis of atherosclerosis and diabetes will be tested. The possible role of self-association in protecting apoA-I from conditions which promote amyloid fibril formation, a contributing mechanism to atherosclerosis progression, will be also evaluated. These studies are highly significant for public health because determining a new functional aspect of apoA- I, which is one of the most important known anti-atherogenic factors, bears potential for the formulation of new therapies and the development of new biomarkers for the evaluation of cardiovascular disease risk.