Mg. Wubbolts et al., TOL PLASMID-SPECIFIED XYLENE OXYGENASE IS A WIDE SUBSTRATE RANGE MONOOXYGENASE CAPABLE OF OLEFIN EPOXIDATION, Enzyme and microbial technology, 16(7), 1994, pp. 608-615
Xylene oxygenase, which is encoded on the TOL plasmid pWWO of Pseudomo
nas putida mt-2, is a key enzyme system in the degradation of toluene
and xylenes by this organism. It was expressed in an Escherichia coli
recombinant strain carrying the xy1MA structural genes. This recombina
nt, which expressed xylene oxygenase from the heat-shock induced lambd
a P-L promoter, was analyzed for its potential as a biocatalytic tool
so as to effect the oxidation of side chains of aromatic hydrocarbons
to the corresponding alcohols. Compounds that were tested as potential
substrates carried different substituents on the aromatic ring at ort
ho, meta, and para positions, relative to the methyl moiety. Products
that accumulated after administration of the aromatic hydrocarbons to
concentrated suspensions of the recombinant were identified by gas cho
matography and mass spectrometry. Toluene derivatives with ortho subst
ituents could not serve as substrates for the biocatalyst, whereas a n
umber of meta- or parasubstituted analogs were efficiently oxidized to
the corresponding benzylalcohols. Bioconversion of the substrates by
resting cells varied from 3 mu mol min(-1) g(-1) cell dry weight for 1
,3,5-trimethylbenzene to 18 mu mol min(-1) g(-1) cell dry weight for m
eta-xylene. Whole cells that expressed xylene oxygenase did catalyze t
he oxidation of the methyl substituent attached to a benzene ring, but
no conversion of n-alkylbenzene derivatives with longer side chains w
as observed. Although the ethyl group of ethylbenzene could not be con
verted by the biocatalyst, cells containing xylene oxygenase were capa
ble of oxidizing the ethylene side group of styrene to produce styrene
epoxide.