Biotransformation of Toluene to o-Cresol in a Two-Phase System by a Solvent Tolerant Bacterium

Abstract

Seventy three toluene-utilizing bacteria were isolated from soil and activated sludge samples. Among them, eleven isolates were highly tolerant to toluene. One of the toluene-tolerant isolates, designated T-57, was identified as P. putida. Strain T-57 showed tolerance to organic solvents whose log P_ow was equal to or greater than 2.3. Genetic analysis revealed that T-57 possesses the toluene dioxygenase operon. The cis-toluene dihydrodiol dehydrogenase gene (todD) mutant of T-57 was constructed by the gene replacement technique. The todD mutant accumulated o-cresol (maximum 1.7 g/l in the aqueous phase) when it was cultivated in the minimal salt medium supplemented with 3% (vol/vol) toluene and 7% (vol/vol) 1-octanol. Thus, T-57 was thought to be a good candidate of the host strain for bioconversion of hydrophobic substrates in the two-phase (organic-aqueous) system.