Semi-rational directed evolution of a Deepsea－derived P450_S18 for Phenazines Construction

  In this study, by using a reported CYP152 family P450_SPα as the probe, P450_S18 was mined from a deep sea-derived bacteria. P450_S18 can directly transform OPD (1) to 2,3-diaminophenazine (2) through C‒N bond construction, in the presence of H₂O₂. We analyzed the possible residues that may be closely related to the binding of OPD (1) in P450_S18, and used alanine scanning to engineer them. All of the mutants exhibited 1.5~3.5-fold increased activity with F292A as the optimal mutant. Phenazines are heterocyclic nitrogenous aromatics with a dibenzopytazine core. They have a wide range of bioactivities, and Cedomon and Shenqinmycin have been approved as biopesticide. The formation of natural phenazines need multiple steps and enzymes, while P450_S18 could directly construct 2,3-diaminophenazine (2) using H₂O₂. Therefore, P450_S18 could be exploited as a tool enzyme in synthesizing phenazine derivatives with good bioactivities.