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Journal of Bacteriology, September 1999, p. 5693-5700, Vol. 181, No. 18
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Involvement of the cis/trans Isomerase
Cti in Solvent Resistance of Pseudomonas putida
DOT-T1E
Frank
Junker and
Juan L.
Ramos*
Department of Plant Biochemistry and
Molecular and Cellular Biology, Estacion Experimental del Zaidin,
Consejo Superior de Investigaciones Cientificas, E-18008 Granada, Spain
Received 8 February 1999/Accepted 23 June 1999
Pseudomonas putida DOT-T1E is a solvent-resistant
strain that is able to grow in the presence of high concentrations of
toluene. We have cloned and sequenced the cti gene of this
strain, which encodes the cis/trans isomerase, termed Cti,
that catalyzes the cis-trans isomerization of esterified
fatty acids in phospholipids, mainly cis-oleic acid
(C16:1,9) and cis-vaccenic acid
(C18:1,11), in response to solvents. To determine the
importance of this cis/trans isomerase for solvent
resistance a Cti-null mutant was generated and characterized. This
mutant showed a longer lag phase when grown with toluene in the vapor
phase; however, after the lag phase the growth rate of the mutant
strain was similar to that of the wild type. The mutant also showed a
significantly lower survival rate when shocked with 0.08% (vol/vol)
toluene. In contrast to the wild-type strain, which grew in liquid
culture medium at temperatures up to 38.5°C, the Cti-null mutant
strain grew significantly slower at temperatures above 37°C. An
in-frame fusion of the Cti protein with the periplasmic alkaline
phosphatase suggests that this constitutively expressed enzyme is
located in the periplasm. Primer extension studies confirmed the
constitutive expression of Cti. Southern blot analysis of total DNA
from various pseudomonads showed that the cti gene is
present in all the tested P. putida strains, including
non-solvent-resistant ones, and in some other Pseudomonas species.
*
Corresponding author. Mailing address: CSIC-Estacion
Experimental del Zaidin, Profesor Albareda 1, E-18008 Granada, Spain. Phone: 34-958-121011. Fax: 34-958-129600. E-mail:
jlramos{at}eez.csic.es.
Journal of Bacteriology, September 1999, p. 5693-5700, Vol. 181, No. 18
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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