Deciphering the Molecular and Functional Basis of RHOGAP Family Proteins: A SYSTEMATIC APPROACH TOWARD SELECTIVE INACTIVATION OF RHO FAMILY PROTEINS
Deciphering the Molecular and Functional Basis of RHOGAP Family Proteins: A SYSTEMATIC APPROACH TOWARD SELECTIVE INACTIVATION OF RHO FAMILY PROTEINS
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DOI：
10.1074/jbc.m116.736967
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发表时间：
2016-09-23
影响因子：
4.8
通讯作者：
Ahmadian, Mohammad R.
中科院分区：
文献类型：
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作者：
Amin, Ehsan;Jaiswal, Mamta;Ahmadian, Mohammad R.
Rho GTPase激活蛋白(RHOGAPs)是Rho相关蛋白家族的主要调节蛋白之一，在许多细胞过程、运动、收缩、生长、分化和发育中起着至关重要的作用。通过数据库搜索，我们提取了66个不同的人类RHOGAP，其中57个具有共同的催化结构域，能够通过刺激缓慢的内在GTP水解酶(GTPase)反应来终止Rho蛋白信号转导。RHOGAP家族大多数成员的特异性在很大程度上尚不清楚。在这里，我们通过结合我们的体外数据和电子计算机数据，全面研究了RHOGAP和Rho蛋白之间的序列-结构-功能关系。实时测定了RHOGAP家族的14个代表对12个Rho家族蛋白的活性。我们确定并从结构上验证了RHOGAP和Rho蛋白界面上的热点是结合和催化的关键决定因素。我们发现，RHOGAP结构域本身是非选择性的，在某些情况下，在无细胞条件下效率相当低。因此，我们认为RHOGAP的其他结构域赋予底物特异性并微调其在细胞中的催化效率。
RHO GTPase-activating proteins (RHOGAPs) are one of the major classes of regulators of the RHO-related protein family that are crucial in many cellular processes, motility, contractility, growth, differentiation, and development. Using database searches, we extracted 66 distinct human RHOGAPs, from which 57 have a common catalytic domain capable of terminating RHO protein signaling by stimulating the slow intrinsic GTP hydrolysis (GTPase) reaction. The specificity of the majority of the members of RHOGAP family is largely uncharacterized. Here, we comprehensively investigated the sequence-structure-function relationship between RHOGAPs and RHO proteins by combining our in vitro data with in silico data. The activity of 14 representatives of the RHOGAP family toward 12 RHO family proteins was determined in real time. We identified and structurally verified hot spots in the interface between RHOGAPs and RHO proteins as critical determinants for binding and catalysis. We have found that the RHOGAP domain itself is nonselective and in some cases rather inefficient under cell-free conditions. Thus, we propose that other domains of RHOGAPs confer substrate specificity and fine-tune their catalytic efficiency in cells.