Inhibition by Methyl 4-nitrophenyl hexylphosphonate 6RB0 gives the same adduct MHH (methyl hydrogen (R)-hexylphosphonate) on the active serine as 4-methylumbelliferyl-hexylphosphonate 3ICW
1 structure: 6RB0: Structure of ester-hydrolase EH1AB1 from the metagenome of lake Arreo complexed with a derivative of methyl 4-nitrophenyl hexylphosphonate
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 pro-teins 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 multifunc-tional metalloenzyme.
