Allosteric effects in the complexes between Ras proteins and Raf

该项目的重点是识别调节细胞增殖的关键蛋白Ras和Raf的分子机制，通过影响它们相互作用细节的天然变体实现不同的生物学功能。该研究建立在早期发现的基础上，其中Ras和Raf之间的相互作用促进了复合物的二聚化，蛋白质的微小变化导致生化结果的变化。该项目挑战了目前与细胞增殖相关的信号传导和调控的教条，并有可能改变对系统如何在分子水平上工作的基本理解。该项目将为在STEM学科中代表性不足的少数群体提供培训。Ras GTP酶、HRas、KRas和NRas通过Ras/Raf/MEK/ERK途径（MAPK途径）通过其中需要Ras二聚化的过程来控制细胞增殖，但机制上尚不清楚。同源GTdR Rap 1A也与Raf相互作用，但其生物学结果取决于特定的Raf亚型。Raf的Ras结合结构域（Raf-RBD）促进Ras二聚化，导致二聚体的两个末端之间的强变构连接（相距85 μ m），并且最近解决了Ras与Raf的RBD和富含半胱氨酸结构域（CRD）两者复合的晶体结构。CRD在单体和二聚体中接触的Ras表面是Ras GTP酶中同种型特异性残基差异的区域，这表明这些差异可能影响MAPK信号传导的机制。该项目将测试这一假设，即Ras和Rap中变构连接的关键区域的残基差异，在其与Raf的复合物中，导致每个GTdR/Raf对特定的变构调节的细化，对调节和信号传导具有影响。这一假设将使用一系列生物物理和生物化学方法进行测试。 该项目由分子和细胞生物科学部的分子生物物理学小组支持。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This project focuses on discerning the molecular mechanisms through which key proteins that regulate cell proliferation, Ras and Raf, attain different biological functions through natural variants that affect details of their interactions with each other. The research is founded on earlier discoveries where the interaction between Ras and Raf promotes dimerization of the complex and small variations in the proteins result in changes in biochemical outcomes. This project challenges current dogma for signaling and regulation associated with cell proliferation, with the potential of transforming fundamental understanding of how the system works at the molecular level. This project will provide training for underrepresented minorities in STEM disciplines. Ras GTPases, HRas, KRas and NRas control cell proliferation via the Ras/Raf/MEK/ERK pathway (MAPK pathway) through a process in which Ras dimerization is required, but not mechanistically understood. The homologous GTPase Rap1A also interacts with Raf, but its biological outcome varies depending on specific Raf isoforms. The Ras Binding Domain of Raf (Raf-RBD) promotes Ras dimerization resulting in strong allosteric linkages between the two extreme ends of the dimer (85 Å apart) and most recently solved the crystal structure of Ras in complex with both RBD and the Cysteine Rich Domain (CRD) of Raf. The surfaces on Ras contacted by the CRD both in the monomer and in the dimer are regions of isoform specific residue differences in the Ras GTPases, suggesting a mechanism through which these differences could impact MAPK signaling. This project will test the hypothesis that residue differences in key regions of allosteric connections in Ras and Rap, across their complexes with Raf lead to refinement of allosteric modulations particular to each GTPase/Raf pair, with implications to regulation and signaling. This hypothesis will be tested using a series of biophysical and biochemical approaches. This project is supported by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.