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. 2003 Dec;69(12):7401-8.
doi: 10.1128/AEM.69.12.7401-7408.2003.

Transporter-mediated uptake of 2-chloro- and 2-hydroxybenzoate by Pseudomonas huttiensis strain D1

A S Yuroff  1 , G Sabat, W J Hickey
Affiliations

Transporter-mediated uptake of 2-chloro- and 2-hydroxybenzoate by Pseudomonas huttiensis strain D1

A S Yuroff et al. Appl Environ Microbiol. 2003 Dec.

Abstract

We investigated the mechanisms of uptake of 2-chlorobenzoate (2-CBa) and 2-hydroxybenzoate (2-HBa) by Pseudomonas huttiensis strain D1. Uptake was monitored by assaying intracellular accumulation of 2-[UL-ring-14C]CBa and 2-[UL-ring-14C]HBa. Uptake of 2-CBa showed substrate saturation kinetics with an apparent Km of 12.7 +/- 2.6 micromoles and a maximum velocity (Vmax) of 9.76 +/- 0.78 nmol min-1 mg of protein-1. Enhanced rates of uptake were induced by growth on 2-CBa and 2-HBa, but not by growth on benzoate or 2,5-di-CBa. Intracellular accumulations of 2-CBa and 2-HBa were 109- and 42-fold greater, respectively, than the extracellular concentrations of these substrates and were indicative of uptake mediated by a transporter rather than driven by substrate catabolism ("metabolic drag"). Results of competitor screening tests indicated that the substrate range of the transporter did not include other o-halobenzoates that serve as growth substrates for strain D1 and for which the metabolism was initiated by the same dioxygenase as 2-CBa and 2-HBa. This suggested that multiple mechanisms for substrate uptake were coupled to the same catabolic enzyme. The preponderance of evidence from tests with metabolic inhibitors and artificial electrochemical gradients suggested that 2-CBa uptake was driven by ATP hydrolysis. If so, the 2-CBa transporter would be the first of the ATP binding cassette type implicated in uptake of haloaromatic acids.

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Figures

FIG. 1.
FIG. 1.
Uptake of 2-CBa and 2-HBa by live and heat-killed cells of P. huttiensis strain D1. Cells used in the experiment were harvested from a continuous culture growing on 2-CBa. Each data point is the mean value from three replicate assays, and error bars indicate the standard deviation of the means.
FIG. 2.
FIG. 2.
Effect of anaerobic conditions and artificially induced electrochemical gradients on 2-CBa uptake. (A) Cells incubated under anaerobic conditions; the arrow indicates the point at which some selected vials were opened to air. (B) Cells with Δp dissipated or reestablished, incubated under aerobic or anaerobic conditions. (C) Dissipation or reestablishment of ΔpH or ΔΨ in cells incubated under aerobic or anaerobic conditions. Each data point in all figures is the mean value of three replicate assays, and error bars indicate the standard deviation of the means (obscured by symbols in some cases). Cells used in the experiment were harvested from a continuous culture growing on 2-CBa.
FIG. 3.
FIG. 3.
Physical map of the hybEFG genes and adjoining regions. Locations of restriction sites for the enzymes listed are indicated by the vertical bars and are numbered (in kilobases) relative to distance from the beginning of the 26-kb region in which they were originally identified (15). For EcoRI, a restriction site located at 10.5 kb that is off the map at the scale used is indicated by the gap. The dashed line indicates the 4-kb region in which the 3.7-kb deletion occurred in the hybFG mutant. Solid lines indicate probes used in Southern hybridization analysis of the mutant.
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References

    1. Alexeyev, M. F., I. N. Shokolenko, and T. P. Croughan. 1995. New mini-Tn5 derivatives for insertion mutagenesis and genetic engineering in Gram-negative bacteria. Can. J. Microbiol. 41:1053-1055. - PubMed
    1. Allende, J. L., A. Gibello, A. Fortun, G. Mengs, E. Ferrer, and M. Martin. 2000. 4-Hydroxybenzoate uptake in an isolated soil Acinetobacter sp. Curr. Microbiol. 40:34-39. - PubMed
    1. Allende, J. L., A. Gibello, A. Fortun, M. Sanchez, and M. Martin. 2002. 4-Hydroxybenzoate uptake in Klebsiella planticola DSZ1 is driven by delta pH. Curr. Microbiol. 44:31-37. - PubMed
    1. Allende, J. L., A. Gibello, M. Martin, and A. Garrido-Pertierra. 1992. Transport of 4-hydroxyphenylacetic acid in Klebsiella pneumoniae. Arch. Biochem. Biophys. 292:583-588. - PubMed
    1. Allende, J. L., M. Suarez, M. Gallego, and A. Garrido-Pertierra. 1993. 4-Hydroxybenzoate uptake in Klebsiella pneumoniae is driven by electrical potential. Arch. Biochem. Biophys. 300:142-147. - PubMed

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