Covalent Fragment Inhibits RhoA Activation by Guanine Exchange Factors.
Covalent Fragment Inhibits RhoA Activation by Guanine Exchange Factors.
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共价片段通过鸟嘌呤交换因子抑制 RhoA 激活。
DOI：
10.1021/acschemneuro.3c00154
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发表时间：
2023
影响因子：
5
通讯作者：
Meroueh,SamyO
中科院分区：
文献类型：
--
作者：
Hussain,MuhammadS;Liu,Degang;Alilain,WarrenJ;Meroueh,SamyO
Ras 同源基因家族成员 (RhoA) 是一种 GTP 酶，也是 GTP 酶 RAS 超家族的成员。 RhoA 是肌动蛋白细胞骨架的主要调节因子。它抑制轴突生长，防止脊髓和脑外伤后的修复和恢复。尽管对 Rho GTP 酶的生物学功能进行了数十年的研究，但仍不存在小分子 Rho 抑制剂。在这里，我们筛选了半胱氨酸亲电子试剂库，以探讨 Cys-107 处的共价键形成是否会导致鸟嘌呤交换因子 Trio 抑制 RhoA 活化。两个片段，丙啶酰胺 1 (ACR-895) 和丙烯酰胺 2 (ACR-917)，以时间依赖性方式抑制 Trio 的 RhoA 核苷酸交换。这些片段与野生型 RhoA 形成共价键，但不与 Cys107Ser RhoA 突变体形成共价键。时间和浓度依赖性研究得出了与个位数小时范围内的 t1/2 值相对应的平衡常数 KI 和反应速率。一个片段对 RhoA 的选择性高于 Rac1 GTPase，并且对 SOS1 的 KRAS 核苷酸交换没有影响。这些片段不抑制 RhoA 与 ROCK 效应蛋白的结合。这项工作确立了 Cys-107 作为 Rho GTPase 抑制的合适位点，并为未来开发 Rho GTPase 共价抑制剂提供了片段起点，这可能对中枢神经系统损伤患者的治疗产生深远的影响。
Ras homolog gene family member (RhoA) is a GTPase and a member of the RAS superfamily of GTPases. RhoA is a master regulator of the actin cytoskeleton. It inhibits axon growth preventing repair and recovery following spinal cord and traumatic brain injuries. Despite decades of research into the biological function of Rho GTPases, there exist no small-molecule Rho inhibitors. Here, we screen a library of cysteine electrophiles to explore whether covalent bond formation at Cys-107 leads to inhibition of RhoA activation by guanine exchange factor Trio. Two fragments, propiolamide1(ACR-895) and acrylamide2(ACR-917), inhibited RhoA nucleotide exchange by Trio in a time-dependent manner. The fragments formed a covalent bond with wild-type RhoA but not Cys107Ser RhoA mutant. Time- and concentration-dependent studies led to equilibrium constantsKIs and reaction rates that correspond to t1/2values in the single-digit hour range. One fragment was selective for RhoA over Rac1 GTPase and had no effect on KRAS nucleotide exchange by SOS1. The fragments did not inhibit RhoA binding to ROCK effector protein. This work establishes Cys-107 as a suitable site for Rho GTPase inhibition and provides fragment starting points for the future development of Rho GTPase covalent inhibitors that could have profound implications in the treatment of patients with injuries of the central nervous system.