Construction of Escherichia coli Strains for Conversion of Nitroacetophenones to ortho -Aminophenols

Abstract

ABSTRACT The predominant bacterial pathway for nitrobenzene (NB) degradation uses an NB nitroreductase and hydroxylaminobenzene (HAB) mutase to form the ring-fission substrate ortho -aminophenol. We tested the hypothesis that constructed strains might accumulate the aminophenols from nitroacetophenones and other nitroaromatic compounds. We constructed a recombinant plasmid carrying NB nitroreductase ( nbzA ) and HAB mutase A ( habA ) genes, both from Pseudomonas pseudoalcaligenes JS45, and expressed the enzymes in Escherichia coli JS995. IPTG (isopropyl-β- d -thiogalactopyranoside)-induced cells of strain JS995 rapidly and stoichiometrically converted NB to 2-aminophenol, 2-nitroacetophenone (2NAP) to 2-amino-3-hydroxyacetophenone (2AHAP), and 3-nitroacetophenone (3NAP) to 3-amino-2-hydroxyacetophenone (3AHAP). We constructed another recombinant plasmid containing the nitroreductase gene ( nfs1 ) from Enterobacter cloacae and habA from strain JS45 and expressed the enzymes in E. coli JS996. Strain JS996 converted NB to 2-aminophenol, 2-nitrotoluene to 2-amino-3-methylphenol, 3-nitrotoluene to 2-amino-4-methylphenol, 4-nitrobiphenyl ether to 4-amino-5-phenoxyphenol, and 1-nitronaphthalene to 2-amino-1-naphthol. In larger-scale biotransformations catalyzed by strain JS995, 75% of the 2NAP transformed was converted to 2AHAP, whereas 3AHAP was produced stoichiometrically from 3NAP. The final yields of the aminophenols after extraction and recovery were >64%. The biocatalytic synthesis of ortho -aminophenols from nitroacetophenones suggests that strain JS995 may be useful in the biocatalytic production of a variety of substituted ortho -aminophenols from the corresponding nitroaromatic compounds.