Exploring the Promiscuity of Phenol Hydroxylase from Pseudomonas stutzeri OX1 for the Biosynthesis of Phenolic Compounds.

Abstract

Enzyme promiscuity plays an important role in developing biosynthetic pathways for novel target products. Phenol hydroxylase (PH) from Pseudomonas stutzeri OX1 is capable of ortho-hydroxylation of phenol and cresol isomers into counterpart catechols. A small ferredoxin-like protein PHQ was clustered together with the ph gene cluster in the genome of P.stutzeri OX1, and its function was not known. In this study, we found that the existence of PHQ has a promotion effect on the catalytic efficiency of PH. Then, we tested the substrate range of PH using nine different non-natural substrates. We found that PH was a promiscuous hydroxylase that could catalyze ortho-hydroxylation of several non-natural substrates, including catechol, 4-hydroxybenzoic acid and resorcinol. On this basis, linking the catechol biosynthetic pathway with the hydroxylation reaction catalyzed by PH enabled construction of a novel biosynthetic pathway for the synthesis of pyrogallol. This work not only characterized a well-performed PH, but also provided a promising hydroxylation platform for the production of high-value phenolic compounds.