Structural and mechanistic studies of INDY proteins

INDY 蛋白的结构和机制研究

基本信息

批准号：
8628114
负责人：
DANENG WANG
金额：
$ 36.87万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-03-01 至 2017-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8628114
关键词：
Adipocytes Adverse effects Affinity Amino Acid Sequence Anions Bacteria Binding Binding Sites Biological Assay Bipolar Disorder Caloric Restriction Cardiovascular Diseases Cations Cell membrane Cells Citrates Coupling Crystallization Dependence Diabetes Mellitus Dimerization Doctor of Philosophy Drug Design Family Fatty Acids Fatty acid glycerol esters Genes Glycolysis Hepatocyte Homologous Gene Human Insulin Resistance Ions Kinetics Knockout Mice Knowledge LDL Cholesterol Lipoproteins Longevity Mammals Measures Membrane Membrane Transport Proteins Molecular Molecular Conformation Molecular Sieve Chromatography Mutagenesis Mutation Obesity Paper Pathway interactions Principal Investigator Proteins Protomer Rattus Reporting Role Site Specificity Structure Structure-Activity Relationship Substrate Specificity Succinates Testing Therapeutic Agents Triglycerides Weight Gain base carboxylate citrate carrier design dicarboxylate dimer energy balance fatty acid biosynthesis fatty acid oxidation fly improved inhibitor/antagonist member nanobodies novel novel strategies programs protein structure public health relevance small molecule sodium ion symporter

项目摘要

DESCRIPTION (provided by applicant): In liver and adipose cells, cytosolic citrate is a major precursor for the synthesis of fatty acids, triacylglycerols, cholesterol and low-density lipoprotein. The cytosolic citrate concentration partially depends on its direct import across the plasma membrane via the Na+-dependent citrate transporter, a member of the divalent anion/Na+ symporter (DASS) family. Mutations of the transporter gene in flies (INDY) result in reduced fat storage through calorie restriction. Knockout mice of the homologous gene are both slimmer and protected from obesity and insulin resistance. Thus, its central role in fatty acid biosynthesis makes NaCT a particularly attractive target of small-molecule therapeutic agents for obesity, diabetes and cardiovascular diseases. We have recently determined the 3.2 ¿ crystal structure of a bacterial INDY homolog in its inward-facing conformation. The crystal structure allows us to propose a detailed transport mechanism for the protein, including substrate specificity, ion specificity, ion- substrate coupling and conformational changes that the protein undergoes to accomplish substrate translocation across the membrane. In the current project, we will test this transport mechanism using mutagenesis and transport assays. We will further characterize the transport mechanism of the protein by determining the structure of the outward-facing conformation of the INDY protein from bacteria and mammals. Understanding of the transport mechanism of these transporters, particularly their substrate and ion specificity, will help in the design of drugs for obesity and diabetes.

描述（由申请人提供）：在肝脏和脂肪细胞中，胞质柠檬酸盐是合成脂肪酸、三酰甘油、胆固醇和低密度脂蛋白的主要前体。胞质柠檬酸盐浓度部分取决于其通过质膜的直接输入。 Na+ 依赖性柠檬酸转运蛋白，果蝇中二价阴离子/Na+ 同向转运蛋白 (DASS) 家族的成员。 (INDY) 通过热量限制减少脂肪储存，同源基因的敲除小鼠既苗条又免受肥胖和胰岛素抵抗，因此，它在脂肪酸生物合成中的核心作用使 NaCT 成为小分子治疗药物特别有吸引力的目标。我们最近确定了针对肥胖、糖尿病和心血管疾病的 3.2 ¿细菌 INDY 同系物的向内构象的晶体结构该晶体结构使我们能够提出蛋白质的详细转运机制，包括底物特异性、离子特异性、离子-底物偶联和构象变化。在当前的项目中，我们将通过确定 INDY 的向外构象的结构来进一步表征蛋白质的转运机制。了解这些转运蛋白的转运机制，特别是它们的底物和离子特异性，将有助于设计治疗肥胖和糖尿病的药物。