Structure of sclerostin protein complexes

硬化蛋白复合物的结构

基本信息

批准号：
8086488
负责人：
RAJIV KUMAR
金额：
$ 21.29万
依托单位：
MAYO CLINIC ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-04-01 至 2013-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8086488
关键词：
Affect Affinity Aging Alkaline Phosphatase American Amino Acids Anabolic Agents Apoptosis Binding Binding Proteins Bone Density C-terminal Caspase Cell Differentiation process Cell Proliferation Cell division Complex Crystallization Crystallography Cystine Data Development Disease Docking Embryo Extracellular Domain Extracellular Matrix Proteins Fibroblasts Fracture Genes Health Human Knowledge LDL-Receptor Related Protein 1 Link Mammalian Cell Mediating Medical Mesenchymal Stem Cells Molecular Molecular Conformation Morbidity - disease rate Mutation Names Osteoblasts Osteoclasts Osteocytes Osteogenesis Osteoporosis Pathway interactions Patients Pharmaceutical Preparations Phenotype Play Postmenopause Proteins Reporting Role Sclerosis Signal Transduction Site Skeleton Solutions Structure Surface Plasmon Resonance Syndrome Teriparatide Therapeutic Van Buchem disease Vascular Endothelial Cell Woman Xenopus angiogenesis base bone bone mass bone morphogenetic protein 6 bone morphogenetic protein receptors bone quality cell motility cyr61 protein design insight lipoprotein receptor related protein 5 men mineralization mutant protein complex receptor receptor binding small molecule

项目摘要

DESCRIPTION (provided by applicant): The objective of this R21 application is to investigate at the atomic level, the mechanism of action of sclerostin, an osteocyte-derived, secreted, cystine-knot protein that inhibits bone formation by examining how sclerostin interacts with proteins that play an essential role in mediating its activity. Because the disruption of sclerostin expression or activity increases bone mass insights developed through our studies may be useful in devising treatments for osteoporosis, a disease affecting 10 million Americans. Bone morphogenetic protein 6 (BMP6), cysteine-rich protein 61 (Cyr61) and low-density-lipoprotein receptor-related protein 5 (LRP5) are key sclerostin-associating proteins reported to mediate sclerostin activity. No structural data exists for these protein complexes. Moreover, preliminary docking studies suggest that the conformations observed in two sclerostin NMR solution-structures must be altered to affect binding to these three proteins. We hypothesize that in contrast to its highly disordered structure in the solution-state, the "loop 2" region of sclerostin (amino acids G86-R109) becomes highly structured when sclerostin binds proteins such as BMP6, Cyr61 and LRP5 whose functions it modulates. If true, sclerostin activity might be antagonized by allosteric modulation and by targeting its interaction site(s), through the use of orally effective, small molecules that would generate new bone in patients. In Aim 1 we will determine the molecular mechanism of sclerostin interaction with BMP6; in Aim 2 we will determine the molecular mechanism of sclerostin interaction with Cyr61 C-terminal domain and in Aim 3 we will identify crystallization conditions for the sclerostin complex with LRP5 extracellular domain. In Aims 1-2, our proposed mechanism for sclerostin interaction will be probed by x-ray crystallography followed by analysis of binding-affinities of structure-guided mutants through the use of surface plasmon resonance. In Aim 3 conditions for crystallizing sclerostin-LRP5 1st ?-propeller complexes will be established. Significance: Osteoporosis is a significant medical health problem associated with fractures and considerable morbidity, prevalent particularly in aging and post-menopausal women. While effective bone anti-resorptive drugs are available for osteoporosis treatment, they have little effect on bone formation. The only anabolic agent available now is teriparatide that must be administered parenterally. Sclerostin is an osteocyte derived protein that inhibits bone formation. Antagonizing its function is a potential therapeutic strategy to increase high quality bone. Our studies will allow an atomic-level understanding of the interface between sclerostin and three protein partners that mediate sclerostin function. We anticipate this knowledge would provide a strong basis for development of antagonistic drugs that can be administrated orally, especially if structural results suggest sclerostin might be amenable to allosteric modulation by small-molecules. PUBLIC HEALTH RELEVANCE: Orally effective drugs that build bone are required for the treatment of osteoporosis. By determining the structures of complexes of sclerostin (a bone inhibitory protein) with other proteins with which it interacts, we will be able to obtain the information that will allow the design of such drugs. This effort will greatly help women and men with osteoporosis and osteoporosis-related fractures.

描述（由申请人提供）：该R21应用的目的是在原子水平上进行研究，Sclerostin的作用机理是一种骨细胞衍生的，分泌的，囊结蛋白的作用机理，通过检查孢子蛋白与蛋白质相互作用，在介导其活性中起着重要作用，从而抑制骨形成骨骼。由于硬化蛋白表达或活性的破坏会增加通过我们的研究发展的骨质量洞察力，这对于设计骨质疏松症的治疗方法可能是有用的，骨质疏松症是一种影响1000万美国人的疾病。骨形态发生蛋白6（BMP6），富含半胱氨酸的蛋白61（CYR61）和低密度脂蛋白受体相关蛋白5（LRP5）是关键的硬化蛋白促蛋白，报道了介导硬化蛋白活性的蛋白质。这些蛋白质复合物不存在结构数据。此外，初步对接研究表明，必须改变在两个硬化素NMR溶液结构中观察到的构象，以影响与这三种蛋白质的结合。我们假设，与溶液状态中的高度无序结构相反，当硬化蛋白结合蛋白（例如BMP6，CYR61和LRP5）功能的蛋白质时，硬化蛋白（氨基酸G86-R109）的“环2”区域变得高度结构。如果是真的，则可以通过使用口服有效的小分子来靶向变构调节，并通过靶向其相互作用位点来拮抗性硬化素活性，从而在患者中产生新的骨骼。在AIM 1中，我们将确定硬化蛋白与BMP6相互作用的分子机制；在AIM 2中，我们将确定硬化蛋白与CYR61 C末端结构域相互作用的分子机制，在AIM 3中，我们将确定与LRP5细胞外域的硬化蛋白复合物的结晶条件。在AIMS 1-2中，我们提出的有关硬化蛋白相互作用的机制将通过X射线晶体学探测，然后通过使用表面等离子体的共振分析结构引导突变体的结合率。在AIM 3条件下，将建立结晶硬化蛋白-LRP5 1st？ - 将建立螺旋桨络合物。意义：骨质疏松症是与骨折和相当大的发病率相关的重大医疗健康问题，尤其是在衰老和绝经后妇女中普遍存在。虽然有效的骨抗敏化药可用于骨质疏松症治疗，但它们对骨骼的影响很小。现在唯一可用的合成代谢代理是teriparatide，必须属于肠胃外施用。硬化蛋白是一种抑制骨形成的骨细胞衍生蛋白。拮抗其功能是增加高质量骨骼的潜在治疗策略。我们的研究将使原子级对硬化蛋白的界面与介导硬化蛋白功能的三个蛋白质伴侣之间的界面有了了解。我们预计，这种知识将为可以口服管理的拮抗药物的开发提供强大的基础，尤其是在结构性结果表明硬化蛋白可能适合小分子的变构调节的情况下。公共卫生相关性：治疗骨质疏松症需要口服有效的骨骼的药物。通过确定硬化蛋白（骨抑制性蛋白）与其他相互作用的蛋白质的结构，我们将能够获得允许设计此类药物的信息。这项工作将极大地帮助男性和男性患有骨质疏松症和骨质疏松相关的骨折。