Inborn Errors of Long Chain Fat Metabolism

描述（由申请人提供）： 线粒体脂肪酸β-氧化传统上被认为是一种产生能量的分解代谢途径，但该途径的中间体可以作为合成其他复合脂质的关键底物。本项目的长期目标是确定脂肪酸氧化蛋白在中间代谢中的作用以及由于先天性缺陷导致的脂肪酸氧化蛋白缺乏的临床影响。第一个资助期的目标是表征LCAD，VLCAD和ACAD 9的功能作用，并探索这些酶在人类和小鼠模型中遗传缺陷的后果。在实现这些目标方面都取得了重大进展。我们的主要研究结果包括突破性的鉴定和部分纯化的多功能脂肪酸氧化复合物含有所有的活动，这一途径与线粒体呼吸链超复合物和鉴定的第一个病人LCAD缺乏症，提出与表面活性剂缺乏症的预测。这项更新申请的目的是研究长链脂肪酸氧化在正常代谢和疾病中的作用。它有三个具体目标。具体目标1是表征多功能脂肪酸氧化复合物的结构及其与线粒体呼吸链的相互作用。我推测，这种接触对于将还原当量从脂肪酸氧化传导到呼吸链是至关重要的， 这一过程降低了线粒体能量产生的效率。具体目标1a是通过首先推导多官能的结构来阐明电子转移的机制。 FAOD复合物使用冷冻电子显微镜。具体目标1b是检查患者确定的突变对VLCAD功能的作用，包括它们对FAOD多功能复合物完整性的影响。具体目标1c是表征VLCAD和LCAD缺陷小鼠模型中脂肪酸氧化复合物的完整性。具体目标2是表征ACAD 9基因中的无效突变和点突变对蛋白质结构和功能的不同影响。我假设ACAD 9在线粒体中作为代谢酶和呼吸链组装/稳定因子具有双重功能。具体目标2a是检查来自缺陷人类患者的细胞中ACAD 9的代谢和兼职功能。具体目的2b是表征ACAD 9敲除小鼠模型中的生化和临床效应范围。具体目标3是进一步表征患者和敲除小鼠模型中的LCAD缺陷，并阐明其在表面活性剂代谢中的功能。具体目标3a是检查野生型和LCAD缺陷型原代肺细胞中的表面活性剂代谢。具体目标3b是描述LCAD缺乏症临床症状的发生率和谱。 公共卫生相关性： 酰基辅酶A脱氢酶是维持体内正常化学平衡的重要酶。我们已经确定了两种新的遗传性疾病，其中一种酶导致肝功能衰竭和肺部疾病。研究这些疾病对于更多地了解其临床表现和治疗非常重要。

DESCRIPTION (provided by applicant): Mitochondrial fatty acid ¿-oxidation is traditionally viewed as an energy-generating, catabolic pathway but intermediates of this pathway can serve as key substrates for synthesis of other complex lipids. The long range objective of this project is to define the role of fatty acid oxidaton proteins in intermediary metabolism and the clinical impact of their deficiency due to inborn errors. The goal of the first funding period was to characterize the functional roles of LCAD, VLCAD, and ACAD9 and to explore the ramifications of genetic deficiencies of these enzymes in humans and mouse models. Significant progress has been made on each of these aims. Our chief findings include the ground breaking identification and partial purification of a multifunctional fatty acid oxidation complex containing all of the activities of this pathway in association with the mitochondrial respiratory chain super complexes and identification of the first patient with LCAD deficiency, presenting as predicted with a surfactant deficiency. The goal of this renewal application is to examine the expanding role of long chain fatty acid oxidation in normal metabolism and disease. It has three specific aims. Specific Aim 1 is to characterize the structure of the multifunctional fatty acid oxidation complex and its interaction with the mitochondrial respiratory chain. I hypothesize that this contact is critical to the channeling of reducing equivalents from fatty acid oxidation to the respiratory chain and that disruption of this process reduces the efficiency of mitochondrial energy generation. Specific Aim 1a is to elucidate the mechanism of electron transfer by first deducing the structure of the multifunctional FAOD complex using cryoelectron microscopy. Specific Aim 1b is to examine the role of patient identified mutations on VLCAD function, including their effect on the integrity of the FAOD multifunctional complex. Specific Aim 1c is to characterize the integrity of the fatty acid oxidatin complex in VLCAD and LCAD deficient mouse models. Specific Aim 2 is to characterize the disparate effects of null and point mutations in the ACAD9 gene on protein structure and function. I hypothesize that ACAD9 serves a duel function in mitochondria as a metabolic enzyme and a respiratory chain assembly/stability factor. Specific Aim 2a is to examine the metabolic and moonlighting functions of ACAD9 in cells from deficient human patients. Specific Aim 2b is to characterize the range of biochemical and clinical effects in an ACAD9 knock out mouse model. Specific Aim 3 is to further characterize LCAD deficiency in patients and a knock out mouse model, and to elucidate its function in surfactant metabolism. Specific Aim 3a is to examine surfactant metabolism in wild type and LCAD deficient primary pneumocytes. Specific Aim 3b is to characterize the incidence and spectrum of clinical symptoms in LCAD deficiency. PUBLIC HEALTH RELEVANCE: The acyl-CoA dehydrogenases are important enzymes in maintaining normal chemical balance in the body. We have identified two new genetic disorder of one of these enzymes that leads to liver failure and lung disease. Studying these disorders is important to learn more about their clinical presentation and treatment.