Effect of Apolipoprotein Structural Adaptability

载脂蛋白结构适应性的影响

基本信息

批准号：
8465891
负责人：
ROBERT O'Mara RYAN
金额：
$ 37.82万
依托单位：
CHILDREN'S HOSPITAL & RES CTR AT OAKLAND
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-06-01 至 2015-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8465891
关键词：
Adopted Affect Age of Onset Agonist Alleles Alzheimer&apos s Disease Amino Acids Apolipoprotein E Apolipoproteins Binding Biological Biological Process Calorimetry Cardiovascular Diseases Cell Culture Techniques Cell Surface Receptors Cholesterol Homeostasis Complex Coronary Arteriosclerosis Culture Media Data Defect Disease Disease susceptibility Experimental Designs Family Family member Generations Goals Health Heteronuclear NMR Homeostasis Isotope Labeling Knowledge LDL-Receptor Related Proteins Lead Ligand Binding Ligands Ligation Lipid Binding Lipids Lipoproteins Low Density Lipoprotein Receptor Low-Density Lipoproteins Maintenance Mediating Metabolic Metabolism Modeling Molecular Molecular Conformation Mutation N-terminal NMR Spectroscopy Nature Neurodegenerative Disorders Pathway interactions Plasma Predisposition Protein Isoforms Proteins Recombinants Regulation Research Resolution Risk Role Shapes Site Structural Models Structure Surface Plasmon Resonance Testing Therapeutic Agents Time Titrations X-Ray Crystallography apolipoprotein E-3 apolipoprotein E-4 base design disorder prevention human subject improved insight interest lipid metabolism member novel strategies particle public health relevance receptor receptor binding research study stable isotope three dimensional structure

项目摘要

DESCRIPTION (provided by applicant): Description The long-term goal of our research is to understand how molecular interactions between lipoproteins and cell surface receptors affect lipid metabolism and disease susceptibility. Knowledge of the metabolic roles of apolipoprotein (apo) E and low-density lipoprotein receptor (LDLR) family members will be combined with available structural information in the design of an experimental strategy to dissect determinants of a productive binding interaction. A novel strategy, termed expressed protein ligation, will be employed to introduce stable isotopes into a specific, predetermined, region of apoE that is essential for LDLR binding. The research to be pursued includes three specific aims. 1) Segmental isotope labeled apoE3-N-temrinal domain, in complex with lipid, will be analyzed by multidimensional heteronuclear NMR spectroscopy in experiments designed to distinguish between alternate structural models of the lipid-bound apoE. 2) The contact sites between apoE and the LDLR will be characterized. It is hypothesized that analysis of segmental isotope labeled apoE3-NT7DMPC in complex with a functional LDLR mini-receptor will yield molecular details of their binding interaction. 3) The interaction of apoE with LDL receptor related protein 6 (LRP6) will be studied. The hypothesis that LDL-A repeats located near the transmembrane spanning sequence of LRP6 are involved in apoE ligand binding will be evaluated. In addition, the postulate that a mutation in LRP6 (R611C), associated with coronary artery disease in human subjects, causes a defect in pH dependent apoE ligand release will be tested. The results of these experiments will extend knowledge of the LDLR family, apoE-mediated lipoprotein metabolism and regulation of plasma lipid homeostasis. Based on the fact that aberrations in the LDLR pathway are positively correlated to onset of cardiovascular disease and apoE manifests isoform-specific susceptibility to neurodegenerative diseases, we anticipate that new knowledge gained from these studies will provide insight into molecular mechanisms that regulate key metabolic processes in health and disease.

描述（由申请人提供）：描述我们研究的长期目标是了解脂蛋白与细胞表面受体之间的分子相互作用如何影响脂质代谢和疾病易感性。在设计实验策略的设计中，将对载脂蛋白（APO）E和低密度脂蛋白受体（LDLR）家族成员的代谢作用的了解将与可用的结构信息结合在一起，以剖析生产性结合相互作用的确定性。一种称为表达蛋白结扎的新型策略将用于将稳定的同位素引入特定的，预定的APOE区域，这对于LDLR结合至关重要。要进行的研究包括三个具体目标。 1）在旨在区分脂质结合apoE的替代结构模型的实验中，将通过多维异核NMR光谱分析具有apoE3-n-链型结构域的节段同位素，与脂质的复合物进行复合。 2）将表征APOE和LDLR之间的接触位点。假设与功能性LDLR微型受体的分析在复合物中被apoE3-NT7DMPC分析将产生其结合相互作用的分子细节。 3）将研究APOE与LDL受体相关蛋白6（LRP6）的相互作用。将评估位于apoE配体结合的LRP6跨膜跨膜序列附近的LDL-A重复序列的假设。此外，假设LRP6（R611c）中与人类受试者中冠状动脉疾病有关的突变会导致pH依赖性APOE配体释放的缺陷。这些实验的结果将扩展对LDLR家族的知识，APOE介导的脂蛋白代谢以及血浆脂质稳态的调节。基于以下事实：LDLR途径中的畸变与心血管疾病的发作呈正相关，而APOE表现出对神经退行性疾病的同工特异性敏感性，我们预计从这些研究中获得的新知识将提供对调节健康和疾病中关键代谢过程的分子机制的洞察力。