Identification and Molecular Characterization of an Efflux Pump Involved in Pseudomonas putida S 12 Solvent Tolerance
Jasper KIEBOOM*a, Jon J. DENNIS*b, Gerben J. ZYSTRA*b, and Jan A.M. de BONT*a
a Division of Industrial Microbiology, Department of Food Technology and Nutrition Sciences, P.O. Box 8129, 6700 EV Wageningen, The Netherlands
b Center for Agricultural Molecular Biology, Foran Hall, 59 Dudley Road, Cook College, Rutgers The State University of New Jersey, New Brunswick NJ 08901-8520, USA
Since the first isolation of a solvent-tolerant Pseudornonas strain by Inoue and Horikoshi in 1989, many different mechanisms have been described that contribute to solvent tolerance. Despite these efforts no comprehensive overview is available to explain the physiological response of microorganisms to toxic organic solvents. Our laboratory has been investigating the ability of P. putida S 12 to withstand toxic concentrations of toluene and other organic solvents. This organism has evolved at least two mechanisms to combat the accumulation of hydrophobic solvents in the membrane. One key observation was the detection of trans rather than cis-unsaturated fatty acids in the membrane of the solvent tolerant bacterium upon exposure to solvents. Recently it has been shown that a second mechanism of solvent tolerance is possessed by P. putida S12. This is an energy-dependent active efflux system for solvents such as toluene that may function in a fashion similar to that for multidrug efflux pumps found in many antibiotic resistant, preserving the integrity of the cell interior. The genes for this solvent efflux pump were cloned from P. putida S 12 and their nucleotide sequence determined. The deduced amino acid sequences encoded by the three genes involved show a striking resemblance to proteins known to be involved in proton-dependent multidrug efflux systems.
