Probing Transcriptional Activation at the Molecular Level

在分子水平上探测转录激活

基本信息

批准号：
8964620
负责人：
ANNA K. MAPP
金额：
$ 33.44万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-04-01 至 2019-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8964620
关键词：
Acetylation Alzheimer&apos s Disease Amino Acids Binding Binding Sites CREB1 gene Cell physiology Cells Chemicals Communication Complement Complex Computer Analysis Data Disease EP300 gene Equilibrium Event Genetic Transcription Goals Health Huntington Disease In Vitro Individual Intervention Kinetics Lead Ligands Malignant Neoplasms Methods Modeling Molecular Molecular Chaperones Molecular Conformation NMR Spectroscopy Neurodegenerative Disorders Neurons Outcome Output Oxidation-Reduction Participant Pathway interactions Pattern Resolution Role Site Structure Supporting Cell Testing Transcription Coactivator Transcription Initiation Transcriptional Activation VP 16 Virus Diseases X-Ray Crystallography analog chemical genetics disulfide bond drug discovery functional outcomes human disease novel painful neuropathy research study small molecule theories therapeutic development therapeutic target tool

项目摘要

DESCRIPTION (provided by applicant): The conformational dynamics of transcriptional activators, coactivators and their complexes underpin regulated transcription. The low energy barriers between individual conformations mean that each participant can use the same group of amino acids to recognize a variety of binding partners, with each complex assuming a distinct conformation. The assumption is that each of the conformations correlates with a unique functional outcome via allosteric communication with other binding partners. However, this model has never been tested, primarily due to a lack of robust tools to connect in vitro observations with function in cells. Our goal is to develop and implement chemical genetic tools for this purpose. Focusing on two functionally important coactivator motifs, KIX (neuropathic pain, neurodegenerative disorders) and AcID (cancer, viral infection), we will develop covalent chemical co-chaperones that stabilize particular conformations of the coactivators alone and in complex with cognate ligands. These co-chaperones will be used to rigorously characterize the structure (X-ray crystallography, NMR spectroscopy) and dynamics (transient kinetics, equilibrium binding, computational analysis) of the coactivators in distinct conformational and assembly states. Complementing the in vitro data will be experiments with chemical co-chaperones in cells, thus identifying both the interactions and the conformations that regulate transcriptional output. Through these studies, allosteric binding sites for small molecule targeting of these disease-relevant transcriptional complexes will be identified.

描述（由申请人提供）：转录激活子、共激活子及其复合物的构象动力学支撑受调控的转录。各个构象之间的低能量势垒意味着每个参与者都可以使用同一组氨基酸来识别各种结合配偶体，每个复合物都呈现不同的构象。假设每个构象通过与其他结合伙伴的变构通讯与独特的功能结果相关。然而，该模型从未经过测试，主要是由于缺乏将体外观察与细胞功能联系起来的强大工具。我们的目标是为此目的开发和实施化学遗传工具。专注于两个功能上重要的共激活子基序，KIX（神经性疼痛，神经退行性疾病）和 AcID（癌症，病毒感染），我们将开发共价化学共伴侣，以稳定单独的共激活子的特定构象以及与同源配体的复合物。这些共伴侣将用于严格表征不同构象和组装状态下共激活剂的结构（X射线晶体学、核磁共振波谱）和动力学（瞬态动力学、平衡结合、计算分析）。体外数据的补充将是细胞中化学共伴侣的实验，从而确定调节转录输出的相互作用和构象。通过这些研究，将确定这些与疾病相关的转录复合物的小分子靶向的变构结合位点。