Understanding the Functional Dynamics of the TokK Enzyme in Carbapenem Biosynthesis via MD Simulations and QM/MM Calculations

Due to the recent surge in antibiotic resistance, developing novel antibiotics is the demand of the time, and thus, a precise understanding of the catalytic mechanisms of enzymes involved in antibiotic biosynthesis becomes crucial. Here, we present a comprehensive investigation into the catalytic mechanism of TokK, a freshly characterized B₁₂-dependent RSMT enzyme that plays an important role in carbapenem biosynthesis. Using MD simulations, we show how the plasticity of the active site facilitates substrate recognition while the quantum mechanics/molecular mechanics calculations provide a detailed mechanistic understanding of the methyl transfer process, elucidating stereochemical preferences. Notably, we demonstrate the indispensable role of Trp215 in orchestrating the proper orientation of the 5'-dA^• radical for efficient substrate methylation, which strongly correlates with the previous findings where the mutation of Trp215 has severely affected the enzyme activity.