Nitroaromatic compounds have broad application space and the development of biosynthetic pathways offers a green chemistry alternative to hazardous industrial synthesis methods. To this point, metabolic engineering efforts toward de novo nitro-product biosynthesis and investigation into nitro-forming enzymes has been limited. Here, we present an integrated heterologous pathway for the production of a stable, non-toxic nitroaromatic product in Escherichia coli. Expression of four exogenous enzymes redirects metabolic flux in the shikimate pathway, including a previously uncharacterized amine oxygenase, to a nitro-containing amino acid with titers above 0.5 mM. Through a fluorescence-based screen, we have identified an amino-acyl tRNA synthetase (AARS)/tRNA pair capable of selective incorporation of the nitro-containing nsAA. Integration of the metabolic pathway with the AARS/tRNA pair enabled biosynthesis and incorporation of a nitro-containing amino acid intro proteins from a supplemented precursor metabolite, and could enable in-situ production of nitro-containing proteins.

De novo biosynthesis and incorporation of a nitroaromatic amino acid

ACS Fall 2021

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