STRUCTURAL-BASED MUTAGENESIS STUDY ON THE LOOP REGIONS OF STAR PROTEIN

基于结构的星蛋白环区诱变研究

• 批准号: 7955482
• 负责人: WALTER L. MILLER
• 金额: $ 0.89万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7955482
• 关键词: Adrenal Glands; Binding; C-terminal; Charge; Chimera organism; Cholesterol; Computer Graphics; Computer Retrieval of Information on Scientific Projects Database; Computer Simulation; Docking; Funding; Grant; Human; Hydrogen Bonding; Hydrophobicity; Imagery; Informatics; Institution; Lipids; Liposomes; Measures; Membrane; Mitochondria; Modeling; Molecular Conformation; Movement; Mutagenesis; Mutate; Play; Proteins; Proteolysis; Reproduction; Research; Research Personnel; Resistance; Resources; Role; Side; Site; Sodium Chloride; Source; Structure; Surface; United States National Institutes of Health; Work; base; biocomputing; biological adaptation to stress; heuristics; in vivo; mitochondrial membrane; mutant; programs; steroid hormone; steroidogenic acute regulatory protein

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The steroidogenic acute regulatory protein (StAR) facilitates the rapid movement of cholesterol into adrenal and gonadal mitochondria, where cholesterol becomes the substrate for the first step in the synthesis of all steroid hormones. StAR works out the mitochondrial membrane required for the stress responses and for reproduction. The C-terminus of StAR is resistant to proteolysis in ther presence of liposomes, implying that this region is associated with the membrane. Crystal structures are available for two proteins closely related to StAR, MLN64 and StARD4, revealing that two loops are adjacent to the C-terminus. Conformational changes of the loops could pivot the C-terminal helix and open the roof to allow cholesterol to bind to the protein. The two loops contain negative charged residues, while the C-terminus is rich is positively charged residues, suggesting that salt bridges and hydrogen bonds may stabilize the structure. The hydrophobicity of the surface surrounding the loops suggests that the loop could be the membrane docking site for lipid loading and unloading. We plan to determine whether the negatively charged residues in the loop regions play a role in substrate binding and membrane association. First, are building a human StAR protein model using the crystal structures of the closely related proteins MLN64 and StARD4, using the "Chimera" program developed in the Computer Graphics Lab. From the model, we will measure the side-chain distances between the negatively charged residues from the loop regions and the positively charged residues from C-terminus. We will then mutate these residue(s) in silico and evaluate their effects on the protein geometry and conformation. Potentially heuristic mutants will then be built and studied in vivo at the bench.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此，可以在其他清晰的条目中表示。列出的机构是 对于中心，这不一定是调查员的机构。 类固醇生成急性调节蛋白（StAR）促进了胆固醇向肾上腺和性腺线粒体的快速运动，胆固醇成为合成所有类固醇激素的第一步的底物。 Star可以解决应力反应和繁殖所需的线粒体膜。 在存在脂质体的情况下，恒星的C末端对蛋白水解具有抵抗力，这意味着该区域与膜有关。 晶体结构可用于与Star，MLN64和Stard4密切相关的两种蛋白质，表明两个环与C端相邻。 环的构象变化可以旋转C末端螺旋并打开屋顶以使胆固醇与蛋白质结合。 这两个环包含负电荷残基，而C末端富含充电的残基，这表明盐桥和氢键可以稳定结构。 回路周围表面的疏水性表明，环可能是用于脂质载荷和卸载的膜对接位点。 我们计划确定环路区域中带负电荷的残基是否在底物结合和膜关联中起作用。 首先，是使用与计算机图形实验室中开发的“嵌合体”程序的密切相关蛋白MLN64和Stard4的晶体结构建立人类恒星蛋白模型的。 从该模型中，我们将测量来自环路区域的负电荷残基与C末端带正电的残基之间的侧链距离。 然后，我们将在硅中突变这些残基，并评估它们对蛋白质几何形状和构象的影响。 然后，可能会在替补席上在体内建造和研究启发式突变体。