Authors: Sandra Alonso, Gerard Santiago, Isabel Cea-Rama, Laura Fernandez-Lopez, Cristina Coscolín, Jan Modregger, Anna K. Ressmann, Mónica Martínez-Martínez, Helena Marrero, Rafael Bargiela, Marcos Pita, Jose L. Gonzalez-Alfonso, Manon L. Briand, David Rojo, Coral Barbas, Francisco J. Plou, Peter N. Golyshin, Patrick Shahgaldian, Julia Sanz-Aparicio, Víctor Guallar & Manuel Ferrer.

Nat Catal 3, 319–328 (2020).
Date: March 2020.
Doi: 10.1038/s41929-019-0394-4

Enzyme engineering has allowed not only the de novo creation of active sites catalysing known biological reactions with rates close to diffusion limits, but also the generation of abiological sites performing new-to-nature reactions. However, the catalytic advantages of engineering multiple active sites into a single protein scaffold are yet to be established. Here, we report on proteins with two active sites of biological and/or abiological origin, for improved natural and non-natural catalysis. The approach increased the catalytic properties, such as enzyme efficiency, substrate scope, stereoselectivity and optimal temperature window, of an esterase containing two biological sites. Then, one of the active sites was metamorphosed into a metal-complex chemocatalytic site for oxidation and Friedel–Crafts alkylation reactions, facilitating synergistic chemo- and biocatalysis in a single protein. The transformations of 1-naphthyl acetate into 1,4-naphthoquinone (conversion approx. 100%) and vinyl crotonate and benzene into 3-phenylbutyric acid (≥83%; e.e. >99.9%) were achieved in one pot with this artificial multifunctional metalloenzyme.