Models of the cooperative mechanism for Rho effector recognition - Implications for RhoA-mediated effector activation
Models of the cooperative mechanism for Rho effector recognition - Implications for RhoA-mediated effector activation
复制标题
DOI：
10.1074/jbc.m409551200
复制
发表时间：
2004-12-17
影响因子：
4.8
通讯作者：
Ahmadian, MR
中科院分区：
文献类型：
--
作者：
Blumenstein, L;Ahmadian, MR
Rho家族的激活的GTP酶调节一系列功能多样的下游效应物，通过效应物蛋白的相互作用和激活启动信号转导途径的网络。尽管效应物被定义为选择性结合小GTP酶的GTP结合状态的蛋白质，但也有几种迹象表明存在核苷酸非依赖性结合模式。通过表征RhoA与其效应子相互作用的分子机制，我们已经确定了三种不同效应子蛋白（Rhotekin，ROCKI/ROKbeta/p160 ROCK，PRK 1/ PKNalpha，其中ROK是RhoA结合激酶）的几个Rho结合结构域对于RhoA的平衡解离常数。GDP和RhoA。GTP荧光光谱法。此外，我们已经确定了两个新的Rho相互作用的结构域在ROCKI，结合RhoA与高亲和力，但不Cdc 42或Rac 1。我们的研究结果，以及最近的结构数据，支持多个效应器结合位点的RhoA的概念，并强烈表明PRK 1和ROCKI的合作结合机制，可能是Rho介导的效应器激活的分子基础。
Activated GTPases of the Rho family regulate a spectrum of functionally diverse downstream effectors, initiating a network of signal transduction pathways by interaction and activation of effector proteins. Although effectors are defined as proteins that selectively bind the GTP-bound state of the small GTPases, there have been also several indications for a nucleotide-independent binding mode. By characterizing the molecular mechanism of RhoA interaction with its effectors, we have determined the equilibrium dissociation constants of several Rho-binding domains of three different effector proteins ( Rhotekin, ROCKI/ROKbeta/p160ROCK, PRK1/ PKNalpha where ROK is RhoA-binding kinase) for both RhoA . GDP and RhoA . GTP using fluorescence spectroscopy. In addition, we have identified two novel Rho-interacting domains in ROCKI, which bind RhoA with high affinity but not Cdc42 or Rac1. Our results, together with recent structural data, support the notion of multiple effector-binding sites in RhoA and strongly indicate a cooperative binding mechanism for PRK1 and ROCKI that may be the molecular basis of Rho-mediated effector activation.