Recent Papers

ABSTRACT

Pseudomonas putida mt-2, harbouring TOL plasmid pWW0, is capable of degrading toluene and a range of di- and tri-alkylbenzenes. In this study, chemostat-grown cells (D = 0.05 h^-1, toluene or m-xylene limitation) of this strain were used to assess the kinetics of the degradation of toluene, m-xylene, p-xylene, and a number of their pathway intermediates. The conversion kinetics for the three hydrocarbons showed significant differences: the maximal conversion rates were rather similar (11-14 mmol.h^-1.[g dry wt]^-1) but the specific affinity (the slope of the v vs s curve near the origin) of the cells for toluene (1300 liters [g dry wt]^-1 .h^-1) was only 5 % and 14 % of those found for m-xylene and p-xylene, respectively. Consumption kinetics of mixtures of the hydrocarbons confirmed that xylenes are strongly preferred over toluene at low substrate concentrations. We also determined the maximum flux rates of pathway intermediates through the various steps of the TOL pathway as far as ring cleavage. Supply of 0.5 mM 3-methylbenzyl alcohol or 3-methylbenzaldehyde to fully-induced cells lead to the transient accumulation of 3-methylbenzoate. Accumulation of the corresponding carboxylic acid (benzoate) was also observed after pulses of benzyl alcohol and benzaldehyde, which are intermediates in toluene catabolism. Analysis of consumption and accumulation rates for the various intermediates showed that the maximal rates at which the initial monooxygenation step and the conversion of the carboxylic acids by toluate 1,2-dioxygenase may occur are 2-3 fold lower than those measured for the two intermediate dehydrogenation steps.