Regulation of mitochondrial metabolism by lysine acylation

赖氨酸酰化调节线粒体代谢

• 批准号: 9304197
• 负责人: ERIC S GOETZMAN
• 金额: $ 39.53万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-15 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9304197
• 关键词: Acetylation; Active Sites; Acyl CoA Dehydrogenases; Acylation; Address; Affect; Binding; Binding Proteins; Biological Assay; Blood; Cardiolipins; Carnitine O-Palmitoyltransferase; Chronic Disease; Citric Acid Cycle; Complex; Defect; Development; Diabetes Mellitus; Diagnosis; Dimensions; Disease; Electron Transport; Electrophoresis; Energy Metabolism; Ensure; Enzymes; Etiology; Fatty Acids; Functional disorder; Funding; Future; Genes; Genetic; Goals; Grant; Heart Diseases; Hereditary Disease; Human; In Vitro; Inborn Errors of Metabolism; Inborn Genetic Diseases; Individual; Inner mitochondrial membrane; Kinetics; Knockout Mice; Knowledge; Lipid Binding; Liver; Long-Chain-Acyl-CoA Dehydrogenase; Lysine; Macromolecular Complexes; Malignant Neoplasms; Manipulative Therapies; Mass Spectrum Analysis; Measures; Membrane; Membrane Proteins; Metabolic; Methods; Mitochondria; Modification; Molecular Models; Molecular Weight; Mouse Protein; Muscle function; Mutagenesis; Myocardium; Obesity; Organ; Pathway interactions; Patients; Pharmacotherapy; Play; Police; Post-Translational Protein Processing; Process; Proteins; Recombinants; Regulation; Research; Respiratory Chain; Role; Sirtuins; Site-Directed Mutagenesis; Testing; Therapeutic; Work; acyl-CoA dehydrogenase; deacylation; enzyme activity; fatty acid metabolism; fatty acid oxidation; gene replacement; heart function; improved; liver function; membrane assembly; mitochondrial dysfunction; mitochondrial metabolism; molecular modeling; mortality; mouse model; novel; novel therapeutics; operation; protein complex

PROJECT ABSTRACT Fatty acid oxidation (FAO) is a critical energy producing pathway in heart, muscle, and liver, among other organs. Inborn errors in genes of the FAO pathway are associated with dysfunction in these organs and a high rate of mortality. Additionally, disruptions in FAO are seen in polygenic diseases such as obesity, diabetes, and cancer. With advances in mass spectrometry profiling of blood metabolites, FAO defects can be readily diagnosed. However, despite 30 years of intensive study, treatment options for modulating FAO in human patients remain limited and ineffective. Knowledge gaps regarding the regulation of FAO enzymes and the functional organization of the FAO pathway within the greater landscape of mitochondrial energy metabolism have limited the development of new therapies. In the previous funding period of this grant, we established reversible lysine post-translational modifications (acetylation, succinylation) as regulators of FAO. We showed that sirtuin enzymes, which deacylate target lysines and restore them to the native state, are important players in maximizing function of the FAO pathway. In the present proposal we hypothesize that lysine acylation regulates FAO enzyme activity, localization to the inner mitochondrial membrane, and the assembly of higher- order metabolic complexes between FAO proteins and the respiratory chain. In Specific Aim 1, we will employ in vitro methods that we pioneered in the previous funding period to identify sirtuin-targeted lysines on the membrane-associated FAO enzymes carnitine palmitoyltransferase-2 (CPT2), mitochondrial trifunctional protein (TFP), and acyl-CoA dehydrogenase-9 (ACAD9). We will perform mutagenesis studies to determine the functional role of each of the sirtuin-targeted lysine residues. In Specific Aim 2 we will investigate physical and functional interactions between the three mitochondrial sirtuins (SIRT3, SIRT4, and SIRT5) and the inner mitochondrial membrane. We hypothesize that the sirtuins police the inner mitochondrial membrane in order to facilitate assembly and operation of higher-order metabolic complexes such as those formed between FAO and the electron transport chain. Finally, Specific Aim 3 will evaluate the effects of lysine acylation on these higher-order complexes using a combination of mouse models and protein complexes assembled in vitro. Understanding the role of the sirtuins in regulating FAO and metabolic supercomplexes will lay the ground work for developing new therapies that manipulate mitochondrial function in human patients with inborn errors of metabolism, as well as those with chronic diseases such as obesity, diabetes, and cancer.

项目摘要 脂肪酸氧化（FAO）是心脏，肌肉和肝脏中产生关键能量的途径 器官。粮农组织途径基因中的先天错误与这些器官的功能障碍有关 死亡率。另外，在肥胖，糖尿病和 癌症。随着血液代谢产物质谱分析的进步，粮农组织缺陷很容易被 诊断。但是，尽管进行了30年的深入研究，但调节人粮农组织的治疗选择 患者仍然有限且无效。关于调节粮农组织和的知识差距 线粒体能量代谢的更大景观中粮农组织途径的功能组织 限制了新疗法的发展。在这笔赠款的上一个资金期间，我们建立了 可逆的赖氨酸后翻译后修饰（乙酰化，琥珀酰化）作为粮农组织的调节剂。我们展示了 脱酰酯靶向赖氨酸并将其恢复到本地状态的Sirtuin酶是重要的参与者 在最大化粮农组织途径的功能中。在本提案中，我们假设赖氨酸酰基化 调节FAO酶活性，定位到内部线粒体膜，并组装较高的 粮农组织蛋白与呼吸链之间的代谢复合物。在特定目标1中，我们将雇用 我们在上一个资金期间开创的体外方法，以识别针对Sirtuin靶向赖氨酸的赖氨酸 与膜相关的粮农组织酶Carnitine Palmitoyltransferase-2（CPT2），线粒体三功能 蛋白质（TFP）和酰基-COA脱氢酶-9（ACAD9）。我们将进行诱变研究以确定 每个靶向Sirtuin的赖氨酸残基的功能作用。在特定目标2中，我们将调查身体 以及三个线粒体Sirtuins（SIRT3，SIRT4和SIRT5）与内部的功能相互作用 线粒体膜。我们假设Sirtuins向内部线粒体膜警察 促进高阶代谢络合物的组装和运行，例如粮农组织之间形成的复合物 和电子传输链。最后，特定的目标3将评估赖氨酸酰基化对这些的影响 使用小鼠模型和体外组装的蛋白质复合物组合的高阶复合物。 了解Sirtuins在调节粮农组织和代谢超复合物中的作用将奠定基础 开发新疗法的工作，这些疗法在人类患者患者患者中操纵线粒体功能 新陈代谢，以及肥胖，糖尿病和癌症等慢性疾病的人。