Chemogenetic engineering of nitrobindin toward an artificial epoxygenase

Abstract

Chemogenetic engineering of metalloproteins emerges as a powerful strategy to generate proteins that catalyze non-natural reactions or convert non-natural substrates. Here, we report on an artificial metalloenzyme (ArM) based on the β-barrel protein nitrobindin (NB) equipped with a manganese protoporphyrin IX catalyst (MnPPIX@NB) for the epoxidation of aromatic alkenes. After exchanging the metal cofactor and two rounds of directed evolution, our ArM shows improved activity (>7-fold increase) with an ee of 20%. The evolution campaign also revealed the importance of the proximal ligand for peroxide activation and subsequent oxygen atom transfer. By utilizing cell adhesion-promoting peptides, a facile strategy is presented to immobilize ArMs on the surface of E. coli cells for on-cell catalysis and chemogenetic engineering of ArMs.