ELUCIDATING THE KINETIC MECHANISM OF SCAFFOLD-MEDIATED MAP KINASE SIGNALING

阐明支架介导的 MAP 激酶信号传导的动力学机制

• 批准号: 8169796
• 负责人: WENDELL A LIM
• 金额: $ 0.18万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-12 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169796
• 关键词: Behavior; Biochemical; Cells; Computer Retrieval of Information on Scientific Projects Database; Environment; Funding; Grant; Individual; Institution; Kinetics; Life; Maps; Mass Spectrum Analysis; Measures; Mediating; Methods; Pathway interactions; Phosphorylation Site; Phosphotransferases; Play; Proteins; Reaction; Regulation; Research; Research Personnel; Resources; Role; Sampling; Scaffolding Protein; Signal Transduction; Signaling Protein; Source; Specificity; Surface; Thermodynamics; United States National Institutes of Health; base; extracellular; physical model; protein complex; research study; response; scaffold; transmission process

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. Living cells interact with and respond to their environment through networks of proteins that transduce extracellular signals into specific biochemical responses inside the cell. Specificity within each pathway is essential for proper regulation of diverse cellular behaviors. Scaffold proteins have been suggested to contribute to specificity by tethering individual signaling proteins together into multi-protein complexes. Emerging evidence suggests that scaffold proteins can also play a more direct role in signal transmission by allosterically promoting specific kinase-kinase reactions and by undergoing conformational changes in response to upstream activators. To construct a physical model for how scaffold proteins promote specific kinase-kinase reactions, we will use a combination of kinetic and biophysical approaches. First, we will measure rate constants for individual kinase-kinase reactions in the presence and absence of scaffolds. Second, we will identify key surfaces and interactions required for scaffold action using crystallographic and biochemical methods. A necessary component of this project is the ability to determine if different protein samples have been phosphorylated, and to identify specific sites of phosphorylation in target proteins. The UCSF mass spectrometry facility will be instrumental in obtaining this information. These experiments will provide kinetic and thermodynamic framework for scaffold-mediated signal transduction, which will provide the basis for understanding how specificity is maintained in cell signaling.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 中心，不一定是研究者的机构。 活细胞通过蛋白质网络与环境相互作用并对其做出反应，这些蛋白质网络将细胞外信号转变成细胞内特定的生化反应。 每个途径的特异性对于正确调节不同的细胞行为至关重要。 支架蛋白被认为通过将单个信号蛋白连接在一起形成多蛋白复合物来促进特异性。 新的证据表明，支架蛋白还可以通过变构促进特定激酶激酶反应以及响应上游激活剂而发生构象变化，从而在信号传递中发挥更直接的作用。 为了构建支架蛋白如何促进特定激酶激酶反应的物理模型，我们将结合动力学和生物物理方法。 首先，我们将测量存在和不存在支架的情况下各个激酶-激酶反应的速率常数。 其次，我们将使用晶体学和生化方法确定支架作用所需的关键表面和相互作用。 该项目的一个必要组成部分是能够确定不同的蛋白质样品是否已被磷酸化，并识别目标蛋白质中磷酸化的特定位点。 加州大学旧金山分校的质谱设施将有助于获取这一信息。 这些实验将为支架介导的信号转导提供动力学和热力学框架，这将为理解如何维持细胞信号传导的特异性提供基础。