Subsequently a pre-reactive binding mode for D-PL was obtained according to molecular docking by inserting D-PL into the predicted structure of D-Lac Figure 1F As a consequence two possible conformati

Subsequently, a pre-reactive binding mode for D-PL was obtained through molecular docking by inserting D-PL into the predicted structure of D-Lac (Figure 1F). As a result, two possible conformations, Config1 and Config2, were identified. These conformations differ in the orientation of the hydroxyl group of D-PL, with Config1 pointing towards H79 and Config2 towards Y215. The two conformations of D-PL were independently simulated through MD simulations, and the binding free energy calculated by MM/GBSA suggested that Config2 was not the most favorable conformation for D-PL binding within the D-Lac active site. The binding free energy of Config1 was slightly lower than that of Config2, approximately 1.32 kcal/mol (Table S1). Therefore, Config1 was assumed to be the reaction conformation of D-PL with D-Lac reaction. Additionally, the binding of D-PL to D-Lac resulted in an "open-close" conformational change in the cap structure domain of D-Lac (Figure 1G). The cap-open conformation allowed the active site to contact the solvent, facilitating substrate binding and product release. In contrast, the cap-closed conformation catalyzed the reaction through a nucleophilic attack