2,4-Dichlorophenoxyacetic acid (2,4-D) utilization by Delftia acidovorans MC1 at alkaline pH and in the presence of dichlorprop is improved by introduction of the tfdK gene.
Growth of Delftia acidovorans MC1 on 2,4-dichlorophenoxyacetic acid (2,4-D) and on racemic 2-(2,4-dichlorophenoxy)propanoic acid ((RS)-2,4-DP) was studied in the perspective of an extension of the strain's degradation capacity at alkaline pH. At pH 6.8 the strain grew on 2,4-D at a maximum rate (mu max) of 0.158 h(-1). The half-maximum rate-associated substrate concentration (Ks) was 45 microM. At pH 8.5 mu max was only 0.05 h(-1) and the substrate affinity was mucher lower than at pH 6.8. The initial attack of 2,4-D was not the limiting step at pH 8.5 as was seen from high dioxygenase activity in cells grown at this pH. High stationary 2,4-D concentrations and the fact that mu max with dichlorprop was around 0.2 h(-1) at both pHs rather pointed at limited 2,4-D uptake at pH 8.5. Introduction of tfdK from D. acidovorans P4a by conjugation, coding for a 2,4-D-specific transporter resulted in improved growth on 2,4-D at pH 8.5 with mu max of 0.147 h(-1) and Ks of 267 microM. Experiments with labeled substrates showed significantly enhanced 2,4-D uptake by the transconjugant TK62. This is taken as an indication of expression of the tfdK gene and proper function of the transporter. The uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP) reduced the influx of 2,4-D. At a concentration of 195 microM 2,4-D, the effect amounted to 90% and 50%, respectively, with TK62 and MC1. Cloning of tfdK also improved the utilization of 2,4-D in the presence of (RS)-2,4-DP. Simultaneous and almost complete degradation of both compounds occurred in TK62 up to D = 0.23 h(-1) at pH 6.8 and up to D = 0.2 h(-1) at pH 8.5. In contrast, MC1 left 2,4-D largely unutilized even at low dilution rates when growing on herbicide mixtures at pH 8.5.
Hoffmann
Doreen
D
Department for Environmental Microbiology, UFZ Centre for Environmental Research, Permoserstr. 15, D-04318, Leipzig, Germany.
Müller
Roland H
RH
D015084:2,4-Dichlorophenoxyacetic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001692:Biological Transport; D020605:Delftia acidovorans / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D005818:Genetic Engineering; D006540:Herbicides / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D006540:Herbicides; D026901:Membrane Transport Proteins; C111945:TfdK protein, Ralstonia eutropha; D015084:2,4-Dichlorophenoxyacetic Acid; C010571:dichlorprop