PDF(Pubmed)
Abstract：
Ras is regulated by a specific guanine nucleotide exchange factor Son of Sevenless (SOS), which facilitates the exchange of inactive, GDP-bound Ras with GTP. The catalytic activity of SOS is also allosterically modulated by an active Ras (Ras-GTP). However, it remains poorly understood how oncogenic Ras mutants interact with SOS and modulate its activity. Here, native ion mobility-mass spectrometry is employed to monitor the assembly of the catalytic domain of SOS (SOScat) with KRas and three cancer-associated mutants (G12C, G13D, and Q61H), leading to the discovery of different molecular assemblies and distinct conformers of SOScat engaging KRas. We also find KRasG13D exhibits high affinity for SOScat and is a potent allosteric modulator of its activity. A structure of the KRasG13D•SOScat complex was determined using cryogenic electron microscopy providing insight into the enhanced affinity of the mutant protein. In addition, we find that KRasG13D-GTP can allosterically increase the nucleotide exchange rate of KRas at the active site more than twofold compared to KRas-GTP. Furthermore, small-molecule Ras•SOS disruptors fail to dissociate KRasG13D•SOScat complexes, underscoring the need for more potent disruptors. Taken together, a better understanding of the interaction between oncogenic Ras mutants and SOS will provide avenues for improved therapeutic interventions.
摘要：
Ras受特定鸟嘌呤核苷酸交换因子Sevenless（SOS）的调节，这促进了不活跃的交换，GTP与GDP挂钩的Ras。SOS的催化活性也由活性Ras(Ras-GTP)变构调节。然而,对于致癌Ras突变体如何与SOS相互作用并调节其活性，我们仍然知之甚少。这里,天然离子迁移-质谱用于监测SOS（SOScat）催化域与KRas和三个癌症相关突变体（G12C，G13D,和Q61H），导致发现与KRas接合的SOScat的不同分子组装体和不同构象体。我们还发现KRasG13D对SOScat具有高亲和力，并且是其活性的有效变构调节剂。使用低温电子显微镜确定KRasG13D•SOScat复合物的结构，从而深入了解突变蛋白的亲和力增强。此外,我们发现，与KRas-GTP相比，KRasG13D-GTP可以变构地增加活性位点处KRas的核苷酸交换速率。此外，小分子Ras•SOS干扰物无法解离KRasG13D•SOScat复合物，强调需要更有效的破坏者。一起来看,更好地了解致癌Ras突变体和SOS之间的相互作用将为改善治疗干预提供途径.
