Theoretical Simulation Research On The Release Mechanism Of Pi In Protein Ras Hydrolysis Reaction

Ras GTP is an active form of Ras protein,which performs a vital function in regulating cell growth and division as a key signaling molecule in cells.The experiments and theoretical simulations have shown that the hydrolysis of Ras GTP involves an intermediate Ras GDP·Pi,which then generates Ras GDP to release Pi（H₂PO₄^-）.Moreover,the interactions of Thr35 residue on Ras with Mg²⁺and the interplays between Pi and Mg²⁺also play crucial roles in the release of Pi.1.In this work,full MM and QM/MM umbrella sampling simulations were carried out to investigate the mechanism of Pi release from the intermediate Ras GDP·Pi·Mg²⁺along the reaction coordinates of aforementioned two key interactions.The results of full MM showed that both HRas and KRas tended to lose the interaction between Thr35and Mg²⁺before Pi was released.However,this phenomenon is not obvious in the results of QM/MM,which may due to the fact that the DFTB3 method does not consider the interaction between residue Qln61 and Pi.Therefore,the following study based on the investigation of the conformational changes of GTP·Mg²⁺to obtain accurate force field parameters for Ras GTP.2.To further investigate the hydrolysis process of Ras GTP,a DP machine learning model was applied combined with XYGJ-OS/6-311+G（d,p）DFT calculations to study the conformational changes of GTP·Mg²⁺from C3 to C2 and obtain an accurate force field model.The results showed that the DP model had good predictions for classical force field configurations but poor predictions for the conformations generated from Dee PMD simulations,and active learning converged slowly.Thus,it is clear that the DP model performs poorly in predicting the conformations of flexible molecules like GTP,and classical force fields remain irreplaceable.Therefore,developing accurate classical force field for GTP·Mg²⁺is also an important research direction.