Efflux of chromate by Pseudomonas aeruginosa cells expressing the ChrA protein.
The ChrA protein of Pseudomonas aeruginosa plasmid pUM505 confers resistance to chromate. Using an in vitro system, we reported [Alvarez, A.H. et al. (1999) J. Bacteriol. 181, 7398-7400] that chromate resistance is based on energy-dependent efflux of chromate. It is shown here that ChrA determines in vivo efflux of 51CrO(4)(2-) as well. Chromate-loaded cell suspensions of P. aeruginosa strain PAO1 harboring recombinant plasmid pEPL1, which expresses the ChrA protein, showed accelerated efflux of 51CrO(4)(2-) as compared to the plasmidless chromate-sensitive derivative. After a 10-min loading, about 40% of 51CrO(4)(2-) was lost from resistant cells in 15 min. Chromate efflux by resistant cells showed a typical saturation kinetics with an apparent K(m) of 82+/-11 microM chromate and a V(max) of 0.133+/-0.009 nmol chromate min(-1) (mg protein)(-1). Oxyanions sulfate and molybdate inhibited chromate efflux in a concentration-dependent fashion, whereas arsenate and ortho-vanadate had no significant effect on chromate release. Inhibition of chromate extrusion by valinomycin, nigericin, and carbonyl cyanide m-chlorophenylhydrazone, but not by oligomycin or dicyclohexylcarbodiimide, indicated that chromate efflux was driven by the membrane potential.
Instituto de Investigaciones Químico-Biológicas, Universidad Michoacana, Edificio B-3, Ciudad Universitaria, 58030 Morelia, Mich., Mexico.
Betzabe E
BE
Pimentel
Rafael
R
Moreno-Sánchez
Carlos
C
Cervantes
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics*; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002840:Chromates / Q000493:pharmacokinetics*; D002860:Chromium Radioisotopes; D024881:Drug Resistance, Bacterial; D004734:Energy Metabolism / Q000187:drug effects; D015964:Gene Expression Regulation, Bacterial; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000235:genetics*; D010087:Oxides / Q000378:metabolism; D010957:Plasmids; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D002840:Chromates; D002860:Chromium Radioisotopes; D008565:Membrane Proteins; D010087:Oxides; C073039:ChrA protein, Bacteria