The Molecular Mechanism of P450-catalyzed Amination of the Pyrrolidine Derivative of Lidocaine: Insights from Multiscale Simulations.

Nitrogen heterocycles are key and prevalent motifs in drugs. Evolved variants of cytochrome P450BM3 (CYP102A1) from Bacillus megaterium employ high-valent oxo-iron(iv) species to catalyze the synthesis of imidazolidine-4-ones via an intramolecular C-H amination. Herein, we use multi-scale simulations, including classical molecular dynamics (MD) simulations, quantum mechanical/molecular mechanical (QM/MM) calculations and QM calculations, to reveal the molecular mechanism of the intramolecular C-H amination of the pyrrolidine derivative of lidocaine bearing cyclic amino moieties catalyzed by the variant RP/FV/EV of P450BM3, which bears five mutations compared to wild type. Our calculations show that overall catalysis includes both the enzymatic transformation in P450 and non-enzymatic transformation in water solution. The enzymatic transformation involves the exclusive hydroxylation of the C-H bond of the pyrrolidine derivative of lidocaine, leading to the hydroxylated intermediate, during which the substrate radical would be bypassed. The following dehydration and C-N coupling reactions are found to be much favored in aqueous situation compared to that in the non-polar protein environment. The present findings expand our understanding of the P450-catalyzed C(sp3)-H amination reaction.