MECHANISM OF GALPHA-13 REGULATED RHOGEF ACTIVITY OF P115RHOGEF

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 P115代表了G12类异源三聚体G蛋白和小GTP酶Rho之间的直接联系。P115(102 KDa)是Galpha-13(Ga13，42 kDa)的负性调控因子和下游效应因子。在其N端，p115含有一个Rhogef-RGS(RgRGS)结构域，其功能是针对Ga13的GTP酶激活蛋白(GAP)。与Ga13的结合也能刺激其对RhoA的鸟嘌呤核苷酸交换活性。P115的全球环境基金活性是由串联的DH和PH结构域实现的，这两个结构域与N-末端的rgRGS结构域通过150个残基连接区分开。测定了rgRGS结构域与Ga13的络合物和与RhoA的络合物中的Dh-PH结构域的晶体结构。我们认为，Ga13与p115的结合导致后者经历了实质性的构象变化，可能是在一个或多个铰链点上的结构域旋转。这种构象变化是刺激环境基金p115对RhoA的活性所必需的。我们建议使用小角X射线散射来确定p115及其与Ga13的络合物的分子包膜。原子分辨结构模型至少可用于组成p115的80%的结构域，并且已经确定了Ga13的结构。了解G蛋白调控p115及其同系物的分子基础将有助于我们深入了解这一特定的调控途径，并加深我们对G蛋白信号转导一般机制的基本理解。

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. p115RhoGEF (p115) represents a direct link between the G12 class heterotrimeric G proteins and the small GTPase Rho. p115 (102 kDa) is both a negative regulator and a downstream effector for Galpha-13 (Ga13, 42 kDa). At its N-terminus, p115 possesses a RhoGEF-RGS (rgRGS) domain that functions as a GTPase activating protein (GAP) towards Ga13. Binding to Ga13 also stimulates its Guanine Nucleotide Exchange (GEF) activity towards RhoA. The GEF activity of p115 is carried out by the tandem DH and PH domains, which are separated from the N-terminal rgRGS domain by a 150 residue linker region. Crystal structures of the complex of the rgRGS domain with Ga13 and the DH-PH domains in complex with RhoA have been determined. We propose that the binding of Ga13 to p115 causes the latter to undergo substantial conformational changes, possibly domain rotation at one or more hinge points. Such conformational changes are required for stimulation of the GEF activity of p115 towards RhoA. We propose to use small angle X-ray scattering to determine the molecular envelope of p115, and that of its complex with Ga13. Atomic-resolution structural models are available for at least 80% of the structural domains that comprise p115, and a structure for the Ga13 has been determined. Understanding the molecular basis by which p115 and its homologs are regulated by G proteins should provide insight into this specific pathway for regulation as well as increase our fundamental understanding of general mechanisms of signaling by G proteins.