Preparative scale production of 3-substituted catechols using a novel monooxygenase from Pseudomonas azelaica HBP 1

Pseudomonas azelaica HBP 1 contains a 2-hydroxybiphenyl 3-monooxygenase (E.C. 1.14.13.44, HbpA) which produces 3-substituted catechols from 2-substituted phenols. The tetrameric enzyme (subunit mass 60 kDa) carries FAD as a prosthetic group, uses NADH as cofactor and is homologous to a number of NADPH dependent flavin containing phenol monooxygenases. HbpA regio-selectively oxidizes 2-substituted phenols and has a broad substrate-range. As ortho-substituent it accepts various alkyl-residues, halogen atoms and substituted phenyl-residues. We have used this monooxygenase for the production of S-substituted catechols, which are useful synthons that are difficult to obtain by chemical means. A recombinant E. coli JM101, containing the hbpA gene, was used for whole-cell biotransformations. Since the phenols as well as the corresponding catechols produced are toxic to cells, a process with a limited feed of starting-material combined with in situ product recovery was used. 3-Phenyl-, 3-chloro-, 3-bromo-, 3-ethyl-, 3-propyl-, 3-i-propyl-, and 3-sec-butyl catechol could thus be produced in gram amounts with space-time yields up to 0.45 g I(-1)h(-1). The products were recovered and further purified by recrystallization. Authenticity of all products, some of which had not been described before, was verified by H-1 NMR, C-13 NMR, IR spectroscopy and GC/MS. (C) 1998 Elsevier Science B.V. All rights reserved.