ABCA1 cholesterol efflux & protein-protein interactions

ABCA1 胆固醇流出

• 批准号: 6922977
• 负责人: MICHAEL Leo FITZGERALD
• 金额: $ 41.48万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6922977
• 关键词: Tangier disease; biotechnology; blood lipoprotein transport; cholesterol; gene mutation; laboratory mouse; laboratory rat; macrophage; mass spectrometry; membrane transport proteins; protein protein interaction; protein purification; protein structure function; proteomics

DESCRIPTION (provided by applicant): Cells efflux cholestrol and phospholids to apolipoproteins such as apoA-l as part of a process that maintains whole body lipid homeostatsis. The physiologic importance of lipid efflux is demonstrated by patients suffering from Tangier disease, a rare genetic condition characterized by peripheral neuropathy, premature cardiovascular disease and an absence of circulating HDL. Genetic mapping studies have associated the Tangier phenotype with mutations in the ATP binding cassette transporter ABCA1 and subsequent studies have shown that ABCA1 plays a rate limiting role in the formation of HDL. In the general population elevated HDL levels are inversely correlated with the incidence of cardiovascular disease, thus therapies that increase ABCA1 activity may help prevent disease progression. The objective of this proposal is to describe key structure-function relations that define the ABCA1 efflux mechanism and what role protein-protein interactions play in determining the activity of ABCA1. This broad aim is focused by the analysis of a naturally occurring Tangier mutation that deletes the last 46 amino acids of the transporter. The deleted amino acids are part of a highly conserved domain, and as shown in this proposal, are essential for ABCA1 efflux activity. Analysis of additional synthetic mutants within the deleted region has defined a novel VFVNFA motif that uniquely identifies a subclass of ABCA transporters. This motif is essential for ABCA1 activity and can act in trans to inhibit efflux activity. A proteomic approach has been developed in which ABCA1 and interesting mutant transporters are affinity purified and the co-purify proteins are analyzed by mass spectrometry. The proteins which interact with the VFVNFA motif are described and their functional relevance to the efflux mechanism investigated will be investigated. Since a set of the interactions appear to down-regulate efflux activity disruption of such interactions may offer therapeutic targets by which ABCA1 efflux can be increased.

描述（由申请人提供）：作为维持全身脂质稳态过程的一部分，细胞将胆固醇和磷脂流出至载脂蛋白（例如apoA-1）。患有丹吉尔病的患者证明了脂质流出的生理重要性，丹吉尔病是一种罕见的遗传性疾病，其特征是周围神经病变、早发心血管疾病和循环 HDL 缺乏。基因图谱研究将 Tangier 表型与 ATP 结合盒转运蛋白 ABCA1 的突变联系起来，随后的研究表明 ABCA1 在 HDL 的形成中起着限速作用。在一般人群中，HDL 水平升高与心血管疾病的发病率呈负相关，因此增加 ABCA1 活性的治疗可能有助于预防疾病进展。该提案的目的是描述定义 ABCA1 外排机制的关键结构-功能关系，以及蛋白质-蛋白质相互作用在确定 ABCA1 活性中发挥的作用。这一广泛的目标是通过分析自然发生的丹吉尔突变来实现的，该突变删除了转运蛋白的最后 46 个氨基酸。删除的氨基酸是高度保守结构域的一部分，如本提案所示，对于 ABCA1 外排活性至关重要。对删除区域内其他合成突变体的分析确定了一个新的 VFVNFA 基序，该基序唯一地识别了 ABCA 转运蛋白的亚类。该基序对于 ABCA1 活性至关重要，并且可以反式作用以抑制外排活性。已经开发出一种蛋白质组学方法，其中 ABCA1 和有趣的突变转运蛋白被亲和纯化，并通过质谱分析共纯化的蛋白质。描述了与 VFVNFA 基序相互作用的蛋白质，并将研究它们与所研究的外排机制的功能相关性。由于一组相互作用似乎下调了外排活性，因此此类相互作用的破坏可能提供增加 ABCA1 外排的治疗靶点。