Epac/cAMP-GEF, A Novel Intracellular cAMP Receptor

CAMP介导的信号转导调节生理条件下许多重要的生物学过程 病情和疾病状态，包括糖尿病、心力衰竭和癌症。在真核细胞中， CAMP由两个普遍表达的细胞内cAMP受体介导，cAMP受体是经典的蛋白激酶 A/cAMP依赖的蛋白激酶(PKA/CAPK)和新近发现的直接激活的交换蛋白 通过cAMP/cAMP调节的鸟核苷酸交换因子(EPAC/cAMP-全环基金)。两个人的存在 无处不在表达的cAMP效应器提供了一种更精确和集成的控制机制 CAMP信号通路在空间和时间上的变化。然而，人们对其发病机制知之甚少。 EPAC激活。这项建议的目标是通过绘制 与EPAC激活相关的构象变化。具体地说，我们已经计划了与 具体目标如下：1)通过定点定位确定EPAC激活所需的特定残基 突变；2)描述与cAMP结合和EPAC激活相关的构象变化 为了确定EPAC及其下游效应子Rap1之间的蛋白质界面，使用增强的 氢交换质谱仪(DXMS)和小角X射线散射；3)解决晶体 用X射线结晶学研究Epac2-cAMP的结构我们研究的长期目标是了解 EPAC的生理功能和调节机制。在此基础上完成建议的研究 应用程序将大大推动该领域朝着这些目标前进。此外，医疗和医疗服务 这一研究项目的药理学意义也是深远的。更好地了解夏令营- 介导的信号转导可能导致新的基于机制的治疗方法的确定 专门针对营地信号组件的战略。

cAMP-mediated signaling regulates a myriad of important biological processes under physiological conditions and disease states, including diabetes, heart failure and cancer. In eukaryotic cells, the effects of cAMP are mediated by two ubiquitously expressed intracellular cAMP receptors, the classic protein kinase A/cAMP-dependent protein kinase (PKA/cAPK) and the recently discovered exchange protein directly activated by cAMP/cAMP-regulated guanine nucleotide exchange factor (Epac/cAMP-GEF). The existence of two ubiquitously expressed cAMP effectors provides a mechanism for a more precise and integrated control of the cAMP signaling pathways in a spatial and temporal manner. However, little is known about the mechanism of Epac activation. The objective of this proposal is to fill the gap in our current knowledge by mapping the conformational changes associated with Epac activation. Specifically, we have planned experiments with the following Specific Aims: 1) To determine the specific residues important for Epac activation by site-directed mutagenesis; 2) to delineate the conformational changes associated with cAMP binding and Epac activation and to determine the protein interface between Epac and it's downstream effector, Rap1, using enhanced deuterium exchange-mass spectrometry (DXMS) and small-angle X-ray scattering; and 3) to solve the crystal structure of Epac2-cAMP using X-ray crystallography. The long-term goals of our research are to understand the physiological functions and mechanisms of Epac regulation. Accomplishing the proposed research in this application will significantly move the field forward towards these goals. Furthermore, the medical and pharmacological implications of this research program are also far-reaching. A better understanding of cAMP- mediated signal transduction could potentially lead to the identification of novel mechanism-based therapeutic strategies specifically targeting the cAMP-signaling components.