Multi-disciplinary Approaches to HDL Structure, Assembly, and Functional Heterogeneity

总结（整个PPG） 新的心脏保护药物和心血管风险的准确指标的开发正在进行 我们认为，由于未能正确识别高密度脂蛋白的心脏保护形式， (HDL)以及对从动脉壁去除胆固醇的机制缺乏足够的了解。这 续期申请将通过继续七个世界级之间的动态互动来解决这些问题。 从不同但互补的观点研究HDL的科学家。高密度脂蛋白，一个重要的调解人， 胆固醇转运是在其最丰富的支架蛋白载脂蛋白A-I（APOA 1） ATP结合盒转运体A1（ABCA 1）。这一过程对于去除胆固醇至关重要。 巨噬细胞事实上，人血浆HDL促进胆固醇流出（CEC）的能力比人血浆HDL促进胆固醇流出（CEC）的能力更强。 诊断心血管疾病（CVD）的风险比传统的测量HDL的胆固醇含量。的 这个多学科计划项目资助的中心假设是，ABCA 1从事高度特异性 与APOA 1颗粒的相互作用可能含有或可能不含有脂质，以产生心脏保护HDL。我们 目的是从分子水平了解这一途径以及HDL亚种在 糖尿病的背景。我们将使用三种通用的重叠方法：1）校准离子迁移率 将特定HDL颗粒的大小和数量与糖尿病患者的CEC和CVD风险相关联， 2)详细的体外机制方法，以阐明允许特定脂化HDL颗粒 与ABCA 1相互作用，以及3）ABCA 1的结构和机制的cryo-EM和计算研究 行动上我们的计划集中在四个地点的三个项目：项目1：超小HDL的生物保护 颗粒- Jay Heinecke，项目负责人; Karin Bornfeldt，Co-I（华盛顿大学）;项目2： ABCA 1介导的胆固醇流出至脂化HDL-W的机制。Sean Davidson，项目负责人 和PPG首席研究员（辛辛那提大学）;项目3： 磷脂/胆固醇易位，ABCA 1- Jere Segrest，项目负责人（范德比尔特大学）; 史蒂夫阿勒，Co-I（亚拉巴马大学伯明翰分校）。此外，四个核心设施将推动科学发展 NIH资源的协同作用和成本效益使用：核心A：行政- W。肖恩·戴维森，核心领导人， 为该计划提供行政支持。核心B：计算生物学- Jere Segrest，核心 负责人，将进行分子建模，同源建模和双/单态正常模式分析 进行结构研究。核心C：脂蛋白定量和功能- Tomas Vaisar，核心负责人; 崇仁汤，Co-I，将量化i）HDL颗粒数量（HDL颗粒的大小和浓度）和 ii）各种HDL和ABCA 1突变体的胆固醇流出能力。核心D：载脂蛋白/脂蛋白生产 - W.核心负责人Sean Davidson将为该计划提供高质量的载脂蛋白， 突变体和脂蛋白样品，两者都是重组衍生的和从人血浆中分离的。

SUMMARY (Entire PPG) The development of new cardioprotective drugs and accurate metrics for cardiovascular risk is being hindered, we believe, by the failure to correctly identify the cardioprotective forms of high-density lipoprotein (HDL) and inadequate knowledge about the mechanisms that remove cholesterol from the artery wall. This renewal application will address these issues by continuing dynamic interactions among seven world-class scientists who study HDL from diverse, but complementary, viewpoints. HDL, an important mediator of cholesterol transport, is created when its most abundant scaffold protein, apolipoprotein A-I (APOA1), interacts with ATP-binding cassette transporter A1 (ABCA1). This process is critical for removing cholesterol from macrophages. Indeed, the capacity for human plasma HDL to promote cholesterol efflux (CEC) is more diagnostic for cardiovascular disease (CVD) risk than the traditional measure of HDL’s cholesterol content. The central hypothesis of this multidisciplinary Program Project Grant is that ABCA1 engages in highly specific interactions with APOA1 particles that may or may not contain lipid to produce cardioprotective HDL. Our objective is to derive a molecular understanding of this pathway and the roles played by HDL subspecies in the setting of diabetes. We will use three general overlapping approaches: 1) calibrated ion mobility spectrometry to relate the size and number of specific HDL particles to CEC and CVD risk in diabetic patients, 2) detailed in vitro mechanistic approaches to unravel the factors that allow specific lipidated HDL particles to interact with ABCA1, and 3) cryo-EM and computational studies of the structure and mechanism of ABCA1’s action. Our plan focuses on three Projects at four sites: Project 1: Cardioprotection by extra-small HDL particles – Jay Heinecke, Project Leader; Karin Bornfeldt, Co-I (University of Washington); Project 2: Mechanism of ABCA1-mediated cholesterol efflux to lipidated HDL – W. Sean Davidson, Project Leader and PPG Principal Investigator (University of Cincinnati); Project 3: Mechanisms of phospholipid/cholesterol translocation by ABCA1 – Jere Segrest, Project Leader (Vanderbilt University); Steve Aller, Co-I (University of Alabama at Birmingham). Additionally, four core facilities will drive scientific synergy and cost-effective use of NIH resources: Core A: Administration – W. Sean Davidson, Core Leader, will provide administrative support for the Program. Core B: Computational Biology – Jere Segrest, Core Leader, will perform molecular modeling, homology modeling, and double/single state normal mode analyses for structural studies. Core C: Lipoprotein Quantitation and Function – Tomas Vaisar, Core Leader; Chongren Tang, Co-I, will quantify i) HDL particle number (the sizes and concentrations of HDL particles) and ii) the cholesterol efflux capacity of various HDLs and ABCA1 mutants. Core D: Apo/Lipoprotein Production – W. Sean Davidson, Core Leader, will supply the Program with high-quality apolipoproteins, customized mutants, and lipoprotein samples, both recombinantly derived and isolated from human plasma.