TGF-beta latency and activation

TGF-β潜伏期和激活

基本信息

批准号：
8963063
负责人：
TIMOTHY A SPRINGER
金额：
$ 39.82万
依托单位：
BOSTON CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-07-07 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8963063
关键词：
Address Affinity Binding Binding Proteins Biochemical Biological Availability Blood Platelets Bone Growth Bone Morphogenetic Proteins Brain Cell surface Cells Cleaved cell Complex Coupled Deuterium Development Disease Drosophila pros protein Endoplasmic Reticulum Extracellular Matrix Family member Fibrosis Glycoproteins Grant Growth Factor Head Health Heart Human Hydrogen Immune Immune response Integrin Binding Integrins Ischemia Kinetics LTBP2 gene Ligands Malignant Neoplasms Marfan Syndrome Mass Spectrum Analysis Measurement Mediating Methods Molecular Molecular Conformation Muscle Muscle Weakness Mutagenesis Mutation Myalgia Peptide Hydrolases Peptides Physiological Play Process Progressive Diaphyseal Dysplasia Protein Binding Protein Isoforms Proteins Proteolysis RGD (sequence)Regulation Regulatory T-Lymphocyte Role Signal Transduction Site Solutions Specificity Stroke Structure Surface Tail Testing Therapeutic Thermodynamics Thrombosis Tissues Transforming Growth Factor beta Wound Healing base bone cell growth dimer in vivo inhibin insight macromolecule monomer mutant nanocage neoplastic cell novel therapeutic intervention novel therapeutics prevent protein complex receptor repaired research study therapeutic development tissue repair tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): Transforming growth factor beta (TGF-ß) plays pivotal roles in development of tissues including bone and muscle, wound healing, immune regulation, and tumor-cell growth and inhibition. TGF-ß regulates repair in all tissues including brain, heart, and bone, and after ischemia and stroke. Mutations in the TGF-ß1 prodomain cause Camurati-Engelmann disease, which manifests as abnormal bone growth with muscle weakness and pain. TGF-ß1, 2 and 3 are synthesized as pro-proteins that dimerize and associate with presenting molecules, latent TGF-ß binding protein (LTBP) or glycoprotein-A repetitions predominant protein (GARP), in the endoplasmic reticulum. Furin cleaves between the prodomain and growth factor domain; however, the prodomain dimer remains noncovalently associated with the growth factor dimer in pro-TGF-ß after secretion. LTBP and GARP bind to the prodomains and store pro-TGF-ßs in the extracellular matrix and on the cell surface, respectively. Although stored in tissues, large latent complexes are biologically inactive because the prodomains encircling the TGF-ß dimer prevent binding to TGF-ß receptors. The key regulatory step in TGF-ß signaling is release of TGF-ß from latency. Integrins aVß6 and aVß8 bind to a specific motif that includes RGD in pro-TGF-ß1 and 3 and activate them, but not pro-TGF-ß2. Integrin-mediated activation requires pro-TGF-ß association with LTBP or GARP. In this grant, we study how aV integrins bind and activate pro-TGF-ß1 and 3, how structural differences between pro-TGF-ß1 and 2 correlate with different activation mechanisms, and how LTBP and GARP contribute to TGF-ß latency. Solving crystal structures of pro-TGF-ß1, integrin aVß6 headpiece alone or in complex with TGF-ß3 peptide, and integrin aVß6 head in complex with human pro-TGF- ß1 macromolecule, coupled with affinity, kinetics, and thermodynamic measurements will reveal conformational changes that occur upon binding and the molecular basis for how integrin aVß6 binds pro-TGF- ß1 with high affinity and specificity, but not pro-TGF-ß2. Potential structure-based therapeutics are also emerging. To better understand TGF-ß structure in the context in which it is kept latent, stored, and activated in vivo, we will study po-TGF-ß1 and 2 and their complexes with LTBP or GARP. We will solve crystal and SAXS structures of pro-TGF-ß1 and 2 and probe their dynamics in solution using proteolysis and hydrogen/deuterium exchange methods. We will solve at least one complex with LTBP or GARP. These experiments will provide key structural insight into conformational changes upon binding. Furthermore, kinetics and affinity measurements on mature TGF-ß1 and 2 binding to free prodomains and LTBP- or GARP-associated prodomains will provide a quantitative, biochemical characterization of their respective interactions. Structural information on pro-TGF-ß1 and 2, and complexes with LTBP or GARP, will provide critical insight into how pro- TGF-ß1 and 2 differ in their latency and activation mechanisms and new therapeutic approaches.

描述（由适用提供）：转化生长因子β（TGF-ß）在骨骼和肌肉，伤口愈合，免疫调节以及肿瘤细胞生长和抑制等组织中起关键作用。 TGF-ß调节所有组织中的修复，包括大脑，心脏和骨骼，以及缺血和中风后。 TGF-ß1prodomain中的突变会引起comurati-engelmann疾病，这表现为异常的骨骼生长，肌肉无力和疼痛。 TGF-ß1、2和3被合成为蛋白质，与呈现分子，潜在的TGF-ß结合蛋白（LTBP）或糖蛋白-a重复的分子相关并相关。术语在prodomain和生长因子结构域之间切割；但是，prodomain二聚体与生长因子二聚体无共价相关分泌后pro-tgf-ß。 LTBP和GARP分别在细胞外基质和细胞表面结合了Prodomains，并将pro-TGF-ß存储在细胞表面上。尽管存储在组织中，但大型潜在复合物在生物学上是无活性的，因为包围TGF-ß二聚体的prodomain会阻止与TGF-ß受体结合。 TGF-ß信号传导中的关键调节步骤是从延迟中释放TGF-β。整合素Avß6和Avß8与特定的基序结合，该基序在Pro-TGF-ß1和3中包括RGD并激活它们，但不激活pro-tgf-ß2。整联蛋白介导的激活需要与LTBP或GARP相关性。在这笔赠款中，我们研究了AV整合素如何结合和激活pro-TGF-ß1和3，即Pro-TGF-ß1和2之间的结构差异如何与不同的激活机制相关，以及LTBP和GARP如何促进TGF-β潜伏期。求解pro-tgf-ß1，整合avß6头台的晶体结构，或与TGF-ß3胡椒的复杂性，以及与人pro-tgf-ß1大分子复杂的整合素avß6头，与亲和力，动力学测量和热力学测量相结合，与构造的构造 - 构造的粘合物变化相结合，与构造的自动化相结合，与构造的构造变化相结合。亲和力和特异性，但不是pro-tgf-ß2。潜在的基于结构的治疗也正在出现。为了更好地理解TGF-ß结构在保持潜在，存储和激活的情况下，我们将研究PO-TGF-β1和2及其与LTBP或GARP的复合物。我们将使用蛋白水解和氢/氘交换方法求解Pro-TGF-β1和2的晶体结构，并在溶液中探测它们的动力学。我们将使用LTBP或GARP解决至少一个复合体。这些实验将为绑定后的会议变化提供关键的结构洞察力。此外，成熟的TGF-ß1和2结合与游离podomains以及LTBP或GARP相关的成果的动力学和亲和力测量将提供其各自相互作用的定量生化特征。 Pro-TGF-ß1和2的结构信息以及使用LTBP或GARP的复合物将对Pro-TGF-ß1和2的潜伏和激活机制以及新的治疗方法有何不同。