CHARACTERIZATION IN-VITRO OF THE HYDROXYLASE COMPONENT OF XYLENE MONOOXYGENASE, THE FIRST ENZYME OF THE TOL-PLASMID-ENCODED PATHWAY FOR THEMINERALIZATION OF TOLUENE AND XYLENES
Jp. Shaw et S. Harayama, CHARACTERIZATION IN-VITRO OF THE HYDROXYLASE COMPONENT OF XYLENE MONOOXYGENASE, THE FIRST ENZYME OF THE TOL-PLASMID-ENCODED PATHWAY FOR THEMINERALIZATION OF TOLUENE AND XYLENES, Journal of fermentation and bioengineering, 79(3), 1995, pp. 195-199
The TOL plasmid from Pseudomonas putida encodes a pathway for the degr
adation of toluene and xylenes. The first step of this degradative pat
hway involves the oxidation of the methyl side chain of the substrates
, which is catalyzed by xylene monooxygenase. Xylene monooxygenase con
sists of two components, an oxygenase component or the XylM protein, a
nd an electron transfer component or the XylA protein. Xylene monooxyg
enase activity was found to be membrane-bound, and Fe2+-dependent. The
activity could be solubilized by two detergents, octyl-beta-glucopyra
noside and olamidopropyl)dimethylammonio]-1-propanesulfonate, After se
paration of the solubilized enzyme by anion exchange chromatography, t
he stability of xylene monooxygenase was drastically reduced. As a con
sequence, the enzyme was characterized using the membrane vesicle frac
tion. The monooxygenase had a pH optimum of 7 and catalyzed the oxidat
ion of toluene, m-xylene, p-xylene and o-xylene, but no activity was o
bserved towards benzyl alcohol.