The Institute of Mathematical Sciences, C.I.T. Campus, Taramani, Chennai, 600113, India.
Homi Bhabha National Institute, Training School Complex, Anushakti Nagar, Mumbai, 400094, India.
Sci Rep. 2018 Aug 28;8(1):12976. doi: 10.1038/s41598-018-31234-7.
The effects of phosphorylation of a serine residue on the structural and dynamic properties of Ras-like protein, Rap, and its interactions with effector protein Ras binding domain (RBD) of Raf kinase, in the presence of GTP, are investigated via molecular dynamics simulations. The simulations show that phosphorylation significantly effects the dynamics of functional loops of Rap which participate in the stability of the complex with effector proteins. The effects of phosphorylation on Rap are significant and detailed conformational analysis suggest that the Rap protein, when phosphorylated and with GTP ligand, samples different conformational space as compared to non-phosphorylated protein. In addition, phosphorylation of SER11 opens up a new cavity in the Rap protein which can be further explored for possible drug interactions. Residue network analysis shows that the phosphorylation of Rap results in a community spanning both Rap and RBD and strongly suggests transmission of allosteric effects of local alterations in Rap to distal regions of RBD, potentially affecting the downstream signalling. Binding free energy calculations suggest that phosphorylation of SER11 residue increases the binding between Rap and Raf corroborating the network analysis results. The increased binding of the Rap-Raf complex can have cascading effects along the signalling pathways where availability of Raf can influence the oncogenic effects of Ras proteins. These simulations underscore the importance of post translational modifications like phosphorylation on the functional dynamics in proteins and can be an alternative to drug-targeting, especially in notoriously undruggable oncoproteins belonging to Ras-like GTPase family.
通过分子动力学模拟研究了丝氨酸残基磷酸化对 Ras 样蛋白 Rap 的结构和动态特性及其与 Raf 激酶 Ras 结合结构域 (RBD) 效应蛋白相互作用的影响，在 GTP 的存在下。模拟表明，磷酸化显著影响 Rap 的功能环的动力学，这些功能环参与与效应蛋白的复合物的稳定性。磷酸化对 Rap 的影响是显著的，详细的构象分析表明，与非磷酸化蛋白相比，磷酸化和 GTP 配体的 Rap 蛋白可以采样不同的构象空间。此外，Rap 蛋白中 SER11 的磷酸化开辟了一个新的腔，可以进一步探索可能的药物相互作用。残基网络分析表明，Rap 的磷酸化导致跨越 Rap 和 RBD 的社区，强烈表明 Rap 中局部改变的变构效应传递到 RBD 的远端区域，可能影响下游信号。结合自由能计算表明，SER11 残基的磷酸化增加了 Rap 和 Raf 之间的结合，这与网络分析结果一致。Rap-Raf 复合物的结合增加可以沿着信号通路产生级联效应，其中 Raf 的可用性可以影响 Ras 蛋白的致癌作用。这些模拟强调了磷酸化等翻译后修饰对蛋白质功能动力学的重要性，并且可以作为药物靶向的替代方法，特别是在属于 Ras 样 GTP 酶家族的难以治疗的致癌蛋白中。
