Structure and mechanism of mammalian stearoyl-CoA desaturases

哺乳动物硬脂酰辅酶A去饱和酶的结构和机制

• 批准号: 10405625
• 负责人: AH-LIM TSAI
• 金额: $ 63.29万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10405625
• 关键词: Acyl Coenzyme A; Address; Anabolism; Binding; Biochemical Reaction; Biological Assay; Cancer Cell Growth; Catalysis; Cell Proliferation; Cells; Coenzyme A; Communication; Complex; Crystallization; Cytochromes b5; Detergents; Diabetes Mellitus; Drug Targeting; Electron Transport; Endoplasmic Reticulum; Enzymes; Family; Fatty acid glycerol esters; Foundations; Freezing; Geometry; Goals; High Fat Diet; Histidine; Homeostasis; Hydrogen; In Vitro; Insecta; Ions; Iron; Kinetics; Knowledge; Length; Life; Ligands; Malignant Neoplasms; Mammalian Cell; Maps; Measures; Mediating; Membrane; Membrane Proteins; Modeling; Molecular Conformation; Movement; Mus; Mutation; NADH; Nitric Oxide; Obesity; Oxidation-Reduction; Oxidoreductase; Oxygen; Palmitoyl Coenzyme A; Pathway interactions; Phospholipids; Positioning Attribute; Problem Solving; Proteins; Reaction; Resolution; Role; Saturated Fatty Acids; Specificity; Spin Trapping; Stearoyl-CoA Desaturase; Structure; System; Testing; Visualization; Zinc; base; cancer cell; cytochrome b5 reductase; diabetic; drug development; fatty acid oxidation; in vitro Assay; insight; lipid biosynthesis; member; nanodisk; novel; obesity treatment; oxidation; pi bond; preference; proteoliposomes; reconstitution; screening; stearoyl-coenzyme A

Mammalian stearoyl-CoA desaturase (SCD) is a member of a super family of redox enzymes that have four transmembrane helices and a diiron center composed of nine conserved histidine residues. This superfamily has over 20,000 members from all kingdoms of life and yet very little is known about the function and mechanism of these proteins. The overall goal of the proposal is to understand the mechanism of SCD at the atomic resolution. SCD resides on endoplasmic reticulum membrane and catalyzes the formation of a double bond on saturated fatty acyl-CoAs. Mice with reduced SCD activity do not become obese or diabetic when fed with a high-fat diet, illustrating the significance of SCD activity in energy homeostasis. SCD expression level is upregulated in cancer cells because of a higher demand for membrane biosynthesis, and inhibition of SCD activity thwarts cancer cell growth. These results led to intense efforts on targeting SCDs for the treatment of obesity, diabetes, and cancer, but the efforts are hampered by a lack of 3-dimenional structures of SCD and incomplete understanding of the catalytic mechanism. It remains unclear how SCDs recognize their substrates and achieve specificity in double bond formation. The all-histidine coordinated diiron center is different from other diiron centers of known structures and its mechanism of catalysis has not been examined. In addition, sustained SCD activity requires two additional membrane proteins, cytochrome b5 (b5) and cytochrome b5 reductase (b5R), which mediate electron transfer from NADH to the diiron center. Yet how the diiron center is redox-cycled through dynamic interactions with b5 and b5R remains a mystery. We expressed and purified functional mouse SCD1 and solved its crystal structure to 2.6 Å resolution. We also expressed and purified full length b5 and b5R, and assembled the enzymatic reactions in vitro. In addition, we produced stable binary complexes between b5R and b5 and between b5 and SCD1, and we established assays to measure electron transfer between the binding partners. These preliminary results allowed us to propose the following three Specific Aims: Aim 1: To characterize the structure and function of SCD1. Aim 2: To understand the oxidative and reductive pathways of the SCD1 diiron center. Aim 3: To understand the structural basis of electron transfer between b5 and SCD1 and between b5R and b5.

哺乳动物硬脂酰辅酶a去饱和酶（SCD）是氧化还原酶超级家族的一员