Characterization and engineering of an o-xylene dioxygenase for biocatalytic applications.

Abstract

The o-xylene dioxygenase from Rhodococcus sp. strain DK17 possesses the unique ability to perform distinct regioselective hydroxylations depending on the size and position of the substituent groups on the aromatic ring. Molecular modeling studies predicted that the substrate-binding pocket of the DK17 o-xylene dioxygenase is large enough to accommodate bicyclics and can be divided into three regions (distal, central, and proximal), and hydrophobic interactions in the distal position are important in substrate binding. Subsequent sitedirected mutagenesis experiments combined with metabolite analysis confirmed these predictions, and further showed that the different positioning of substrates in the active site of the enzyme results in the generation of different products. This review specifically summarizes molecular functional mechanisms that enable this enzyme to catalyze the respective hydroxylation reactions.tudies predicted that the substrate-binding pocket of the DK17 o-xylene dioxygenase is large enough to accommodate bicyclics and can be divided into three regions (distal, central, and proximal), and hydrophobic interactions in the distal position are important in substrate binding. Subsequent sitedirected mutagenesis experiments combined with metabolite analysis confirmed these predictions, and further showed that the different positioning of substrates in the active site of the enzyme results in the generation of different products. This review specifically summarizes molecular functional mechanisms that enable this enzyme to catalyze the respective hydroxylation reactions.