To validate the above-proposed mechanism, we performed QM/MM calculation for the most populated snapshot 1 of the MD simulation. To obtain the reaction profile, we performed the potential energy surface (PES) scan for the H-abstraction of a QM/MM optimized geometry. As can be seen, the Mg 2+ ion is penta-coordinated with three phosphate oxygen and two oxygen of the E130. The ε-Nitrogen of the Histidine is very close (1.8Å) to the OH group of the homoserine, and it is in the optimum position to abstract a proton once initiated. During the PES scan, we found that the γ-Phosphate group easily detaches with ATP, and at the same time, the proton of the substrate shifts to the H138 and forms an intermediate IM. This concerted reaction occurs via a TS energy barrier of 18.25 kcal/mol within the enzymatic range. In the subsequent step, Ob of the γ-Phosphate of the newly formed phosphorylated L-homoserine abstract a proton from the H138 without any barrier and forms the ultimate product (PC) O-phospho-L-homoserine (see Figure ). In a previous study, Krishna et al. proposed a different pathway via a direct proton transfer from the hydroxyl group of the substrate to the non-bridging O atom of the terminal phosphate of ATP followed by phosphate transfer to the substrate. Therefore, we also performed the QM/MM calculation for this possibility. The reaction profile by the PES scan for this pathway is shown in Figure . As can be seen, the activation energy required for this profile is 31.16 kcal/mol, which is far higher than the reaction pathway via H138.