Structure and Function of Enzymes in Fatty Acid Oxidation

脂肪酸氧化酶的结构和功能

• 批准号: 8372063
• 负责人: JUNG JA P. KIM
• 金额: $ 38.25万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1982
• 资助国家: 美国
• 起止时间: 1982-03-01 至 2016-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8372063
• 关键词: Acetyl Coenzyme A; Active Sites; Acyl CoA Dehydrogenases; Acyl Coenzyme A; Affect; Agonist; Amino Acids; Binding; Biochemical; Cardiomyopathies; Cells; Chemicals; Child; Choline; Cleaved cell; Clinical; Coenzyme A; Complex; Crystallization; Development; Diabetes Mellitus; Diet; Disease; Electron Spin Resonance Spectroscopy; Electron Transport; Electron transfer flavoprotein; Electrons; Enoyl-CoA Hydratase; Enzymes; Escherichia coli; Etiology; Exercise; Failure to Thrive; Family; Fasting; Fatty Acids; Flavins; Flavoproteins; Goals; Health; Heart; Hepatocyte; Human; Human Activities; Human body; Inborn Genetic Diseases; Inherited; Inner mitochondrial membrane; Investigation; Kidney; Left; Ligand Binding; Ligands; Light; Liver; Long-Chain-Acyl-CoA Dehydrogenase; Mass Spectrum Analysis; Membrane; Metabolic; Metabolic Diseases; Metabolic syndrome; Metabolism; Methods; Mitochondria; Mitochondrial Diseases; Molecular Conformation; Movement; Multienzyme Complexes; Muscle; Muscle Cells; Mutation; Myocardium; Myopathy; NADP; Neonatal Screening; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Oxidants; Oxidoreductase; Physiological; Play; Pregnancy; Process; Protein Binding; Proteins; Reaction; Recombinant Proteins; Resolution; Respiratory Chain; Role; Series; Structure; Sudden infant death syndrome; System; Testing; Therapeutic; Time; Very Long Chain Fatty Acid; X-Ray Crystallography; acyl-CoA dehydrogenase; base; clinical phenotype; crosslink; dehydrogenation; design; diagnosis design; dimethylglycine dehydrogenase; electron donor; electron-transferring-flavoprotein dehydrogenase; fatty acid oxidation; feeding; heart cell; improved; inhibitor/antagonist; insight; long chain fatty acid; member; mitochondrial membrane; novel; oxidation; polypeptide; protein protein interaction; skeletal; thioester; young adult

DESCRIPTION (provided by applicant): Fatty acid ?-oxidation is the major energy-producing process in the liver, heart, and muscle. It is carried out by a series of four reactions that successively cleave acetyl-CoA from fatty acyl-CoA. The rate of this process can be altered by diet (fed/fasting), physiological status (pregnancy), or diseases (diabetes). The first of the four reactions in this process is initiated by a family of flavoproteins, acyl-CoA dehydrogenases (ADs). Electron transfer from ADs to the mitochondrial OXPHOS chain is catalyzed by electron transfer flavoprotein (ETF) and the membrane-bound ETF-ubiquinoneoxidoreductase (ETF-QO). There are at least seven soluble ADs for catalyzing short chain acyl-CoAs, and two membrane-bound ADs specific for long chain fatty acyl-CoAs, very long chain AD (VLCAD) and ACAD9. The three remaining reactions of ?- oxidation for long chain fatty acids are carried out by the trifunctional protein (TFP), a membrane-bound multienzyme complex. Inborn errors of fatty acid oxidation have emerged as an increasing health problem and now represent the most common group of disorders identified through expanded newborn screening, affecting 2-3/1,000 babies born nationwide. These disorders present sudden infant death syndrome, cause cardiomyopathy, and are the most common cause of skeletal myopathy in older children and young adults. Recently ACAD9 has been shown to be essential for the assembly of Complex I, the largest and most complicated enzyme (~980 kDa with 45 subunits) among the five OXPHOS complexes. Very little is known concerning the mechanism of the assembly process of this important enzyme. Disorders of the mitochondrial OXPHOS system are the most common of inborn metabolic diseases, resulting in a wide variety of clinical phenotypes ranging from exercise intolerance to failure to thrive. We have determined the crystal structures of all but one of the soluble ADs, as well as one membrane-bound AD (VLCAD), ETF, and ETF-QO. The proposed investigations are focused on three membrane-bound enzymes, VLCAD, ACAD9, and TFP, and interactions of ETF with its electron transfer partners, including ADs, dimethylglycine dehydrogenase (DD), and ETF-QO. DD functions in choline metabolism and is not a member of the AD family, but donates electrons to ETF. Specific Aims are: 1) Structural studies of human TFP by X-ray crystallography to understand how its three distinct active sites communicate with each other; 2) Studies of VLCAD, including a) studies of clinical mutations, and b) to determine the orientation of VLCAD on the mitochondrial membrane and interactions with TFP and ETF by EPR spectroscopy; 3) studies of ACAD9 to determine the biochemical/structural basis for its unique role in mitochondrial Complex I assembly; and 4) to investigate the domain movement of ETF and its interactions with three representative electron donors (medium chain acyl-CoA dehydrogenase, VLCAD, and DD) and with its electron acceptor, ETF-QO. PUBLIC HEALTH RELEVANCE: All cells in the human body need energy to maintain their structural integrity and to perform their vital functions. The major proportion of this energy comes from the oxidation of fatty acids in heart, liver, and muscle cells and an imbalance in this process results in disease states, such as obesity and diabetes. A better understanding of how these enzymes function will provide insight into diagnosis and the design and development of novel inhibitors or agonists for the treatment of metabolic disorders.

描述（由申请人提供）：脂肪酸α-氧化是肝脏、心脏和肌肉中主要的能量产生过程。它是通过一系列四个反应来进行的，这些反应依次将乙酰辅酶A从脂肪酰辅酶A上裂解下来。这一过程的速率可能会因饮食（进食/禁食）、生理状态（怀孕）或疾病（糖尿病）而改变。四个中的第一个 这个过程中的反应是由黄素蛋白家族酰基辅酶A脱氢酶（AD）引发的。从 AD 到线粒体 OXPHOS 链的电子转移由电子转移黄素蛋白 (ETF) 和膜结合 ETF-泛醌氧化还原酶 (ETF-QO) 催化。至少有七种用于催化短链酰基辅酶A的可溶性AD，以及两种特异于长链脂肪酰基辅酶A的膜结合AD，即极长链AD（VLCAD）和ACAD9。长链脂肪酸的 β-氧化的其余三个反应由三功能蛋白（TFP）（一种膜结合多酶复合物）进行。先天性脂肪酸氧化错误已成为一个日益严重的健康问题，目前是通过扩大新生儿筛查发现的最常见疾病组，影响全国出生的婴儿的千分之二至三。这些疾病会导致婴儿猝死综合症，导致心肌病，并且是年龄较大的儿童和年轻人骨骼肌病的最常见原因。 最近，ACAD9 已被证明对于复合物 I 的组装至关重要，复合物 I 是五种 OXPHOS 复合物中最大、最复杂的酶（约 980 kDa，有 45 个亚基）。关于这种重要酶的组装过程的机制知之甚少。线粒体 OXPHOS 系统疾病是最常见的先天性代谢疾病，可导致多种临床表型，从运动不耐受到发育迟缓。 我们已经确定了除一种可溶性 AD 之外的所有 AD 以及一种膜结合 AD (VLCAD)、ETF 和 ETF-QO 的晶体结构。拟议的研究重点是三种膜结合酶，VLCAD、ACAD9 和 TFP，以及 ETF 与其电子转移伙伴的相互作用，包括 AD、二甲基甘氨酸脱氢酶 (DD) 和 ETF-QO。 DD 在胆碱代谢中发挥作用，不是 AD 家族的成员，但向 ETF 提供电子。具体目标是： 1) 通过 X 射线晶体学对人类 TFP 进行结构研究，以了解其三个不同的活性位点如何相互通信； 2) VLCAD的研究，包括a)临床突变的研究，b)通过EPR光谱确定VLCAD在线粒体膜上的方向以及与TFP和ETF的相互作用； 3) 研究ACAD9以确定其在线粒体复合物I组装中独特作用的生化/结构基础； 4) 研究 ETF 的结构域移动及其与三种代表性电子供体（中链酰基辅酶 A 脱氢酶、VLCAD 和 DD）及其电子受体 ETF-QO 的相互作用。 公共健康相关性：人体中的所有细胞都需要能量来维持其结构完整性并执行其重要功能。这种能量的主要部分来自心脏、肝脏和肌肉细胞中脂肪酸的氧化，这一过程的不平衡会导致肥胖和糖尿病等疾病。更好地了解这些酶的功能将为诊断以及设计和开发用于治疗代谢紊乱的新型抑制剂或激动剂提供见解。