Molecular basis of fatty acid transport by peroxisomal ABC transporters

过氧化物酶体 ABC 转运蛋白转运脂肪酸的分子基础

• 批准号: 10796495
• 负责人: Amer Alam
• 金额: $ 17.5万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10796495
• 关键词: ACBD3 gene; ATP-Binding Cassette Transporters; Acceleration; Adrenoleukodystrophy; Bile Acid Biosynthesis Pathway; Bile Acids; Biochemical; Biological; Cells; Chemicals; Cryoelectron Microscopy; Defect; Development; Diagnostic; Disease; Disease Pathway; Electron Spin Resonance Spectroscopy; Environment; Family; Fatty Acids; Functional disorder; Future; Impairment; In Vitro; Lipids; Liver diseases; Metabolic; Metabolic Diseases; Molecular; Molecular Conformation; Mutation; Neurologic; Pathology; Patient-Focused Outcomes; Peroxisomal Disorders; Phospholipids; Physiological; Property; Regulation; Resolution; Role; Structure; Substrate Specificity; Therapeutic; Very Long Chain Fatty Acid; Work; branched chain fatty acid; combat; design; drug development; fatty acid metabolism; fatty acid transport; functional disability; improved; in silico; in vitro Assay; insight; leukodystrophy; nervous system disorder; peroxisome; rare genetic disorder; stem; tool

Project Summary Peroxisomal ABC transporters like ABCD1, ABCD2, and ABCD3 shuttle very long chain fatty acids (VLCFAs), branched chain fatty acids (BCFAs), and bile acid precursors into peroxisomes. Their functional impairment leads to severe neurological and metabolic pathologies stemming from disrupted phospholipid and fatty acid metabolism, including X-linked Adrenoleukodystrophy (X-ALD), both the most common leukodystrophy and most common peroxisomal disorder that is caused by mutations in ABCD1 and for which no cure exists, and bile acid synthesis defects and liver disease stemming from impaired ABCD3 function. While additional roles for ABCD transporters in a wider array of disease pathways continue to be uncovered, the underlying mechanisms governing their substrate recognition, transport, and transport regulation remain poorly understood. The long- term objectives of this project are to gain insight into peroxisomal ABCD transporter function and regulation in molecular detail. We will use a combination of biochemical and cell biological tools, high resolution structural analysis by cryo-electron microscopy, and continuous wave electron paramagnetic resonance (CW-EPR) spectroscopy to reveal the functionally relevant structural features and conformational states used by ABCD transporters in fatty acid translocation, how they may be altered by ABCD1 mutation in X-ALD, and how ABCD1 is mechanistically distinct from ABCD2 and ABCD3 despite their functional overlap. Specific Aim 1 deals with the development and utilization of in vitro assays for determining substrate specificity profiles and transport properties of ABCD1, ABCD2, and ABCD3. Specific Aim 2 deals with obtaining high resolution structural information of ABCD1, ABCD2, and ABCD3 in functionally relevant states in a physiological lipid environment through cryo-EM analysis. Specific Aim 3 deals obtaining information on the structural dynamics of ABCD1 through CW-EPR studies. Our results will provide fundamental insights into peroxisomal ABC transporter functioning that can be exploited for ABCD1 targeted diagnostic and therapeutic tools to improve X-ALD patient outcomes, provide a framework for the design and development of chemical probes to study ABCD family function, and generate reliable in vitro and in silico tools to accelerate drug development/discovery efforts targeting them.

项目概要 过氧化物酶体 ABC 转运蛋白如 ABCD1、ABCD2 和 ABCD3 穿梭极长链脂肪酸 (VLCFA)， 支链脂肪酸（BCFA）和胆汁酸前体转化为过氧化物酶体。他们的功能障碍导致 因磷脂和脂肪酸破坏而引起的严重神经和代谢病变 代谢障碍，包括 X 连锁肾上腺脑白质营养不良 (X-ALD)，这既是最常见的脑白质营养不良，也是最常见的脑白质营养不良 由 ABCD1 突变引起且无法治愈的常见过氧化物酶体疾病，以及胆汁酸 ABCD3 功能受损导致合成缺陷和肝病。而 ABCD 的附加角色 更广泛的疾病途径中的转运蛋白不断被发现，其潜在机制 其底物识别、运输和运输调节的控制仍然知之甚少。长- 该项目的长期目标是深入了解过氧化物酶体 ABCD 转运蛋白的功能和调节 分子细节。我们将结合使用生化和细胞生物学工具、高分辨率结构 通过冷冻电子显微镜和连续波电子顺磁共振 (CW-EPR) 进行分析 光谱揭示 ABCD 使用的功能相关结构特征和构象状态 脂肪酸易位中的转运蛋白，X-ALD 中 ABCD1 突变如何改变它们，以及 ABCD1 如何改变 尽管功能重叠，但在机制上与 ABCD2 和 ABCD3 不同。具体目标 1 涉及 开发和利用体外测定来确定底物特异性概况和运输 ABCD1、ABCD2 和 ABCD3 的性质。具体目标 2 涉及获得高分辨率结构 ABCD1、ABCD2 和 ABCD3 在生理脂质环境中处于功能相关状态的信息 通过冷冻电镜分析。具体目标 3 涉及获取 ABCD1 的结构动力学信息 通过 CW-EPR 研究。我们的结果将为过氧化物酶体 ABC 转运蛋白提供基本见解 可用于 ABCD1 靶向诊断和治疗工具以改善 X-ALD 患者的功能 结果，为研究 ABCD 家族的化学探针的设计和开发提供框架 功能，并生成可靠的体外和计算机工具来加速药物开发/发现工作 针对他们。