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: 10700981
• 负责人: Amer Alam
• 金额: $ 37.42万
• 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: 2022-09-10 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10700981
• 关键词: ACBD3 gene; ATP Hydrolysis; ATP-Binding Cassette Transporters; Acceleration; Adrenoleukodystrophy; Bile Acid Biosynthesis Pathway; Bile Acids; Binding; Biochemical; Biological; Biological Assay; Catabolism; Cells; Chemicals; Coenzyme A; Cryoelectron Microscopy; Cysteine; Data; Defect; Development; Diagnostic; Disease; Disease Pathway; Electron Spin Resonance Spectroscopy; Elements; Environment; Esterification; Esters; Family; Fatty Acids; Functional disorder; Future; Genes; Human; Impairment; In Vitro; Label; Length; Link; Lipid Bilayers; Lipid Chemistry; Lipids; Liposomes; Liver diseases; Location; Measures; Mediating; Membrane; Membrane Transport Proteins; Metabolic; Metabolic Diseases; Molecular; Molecular Conformation; Mutation; Neurologic; Nucleotides; Pathologic; Pathology; Pathway interactions; Patient-Focused Outcomes; Pattern; Peroxisomal Disorders; Phenotype; Phospholipids; Physiological; Production; Property; Proteins; Regulation; Resolution; Role; Scanning; Shapes; Specificity; Spin Labels; Structure; Substrate Specificity; Therapeutic; Transmembrane Transport; Very Long Chain Fatty Acid; Work; branched chain fatty acid; combat; design; drug development; fatty acid metabolism; fatty acid transport; functional disability; hydrophilicity; improved; in silico; in vitro Assay; insight; leukodystrophy; mutant; nanodisk; nervous system disorder; peroxisome; proteoliposomes; rare genetic disorder; reconstitution; small molecule; stem; structural determinants; 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.

项目摘要 过氧化物酶体ABC转运蛋白（例如ABCD1，ABCD2和ABCD3班车）非常长的链脂肪酸（VLCFAS）， 分支链脂肪酸（BCFA）和胆汁酸前体成过氧化物酶体。他们的功能障碍引线 因磷脂和脂肪酸干扰而导致的严重神经和代谢病理 新陈代谢，包括X连锁的肾上腺素繁殖（X-ald），既是最常见的白细胞营养不良，又是大多数 由ABCD1突变引起的常见过氧化物酶体疾病，并且不存在治疗，胆汁酸会引起 ABCD3功能受损的合成缺陷和肝病。而ABCD的其他角色 在更广泛的疾病途径中继续发现的转运蛋白，潜在的机制 管理其底物认可，运输和运输法规的理解仍然很差。长期 该项目的术语目标是洞悉过氧化物酶体ABCD转运蛋白函数和调节 分子细节。我们将结合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）分析 光谱法揭示了ABCD使用的功能相关的结构特征和构象状态 脂肪酸易位的转运蛋白，如何通过X-Ald中的ABCD1突变改变它们以及ABCD1的方式 尽管其功能重叠，但在机械上与ABCD2和ABCD3不同。特定目标1处理 用于确定底物特异性曲线和运输的体外测定法的开发和利用 ABCD1，ABCD2和ABCD3的属性。具体目标2与获得高分辨率结构的交易 生理脂质环境中功能相关状态中ABCD1，ABCD2和ABCD3的信息 通过冷冻EM分析。具体目标3交易获取有关ABCD1结构动力学的信息 通过CW-EPR研究。我们的结果将提供对过氧化物酶体ABC转运蛋白的基本见解 可以为ABCD1靶向诊断和治疗工具而利用的功能来改善X-ALD患者 结果，为研究ABCD家族的化学探针设计和开发提供了一个框架 功能，并生成可靠的体外和有机硅工具，以加速药物开发/发现工作 针对它们。