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Applied and Environmental Microbiology, June 1999, p. 2631-2635, Vol. 65, No. 6
Division of Industrial Microbiology,
Department of Food Technology and Nutritional Sciences, Wageningen
Agricultural University, Wageningen, The Netherlands
Received 14 December 1998/Accepted 23 March 1999
Solvent-tolerant microorganisms are useful in biotransformations
with whole cells in two-phase solvent-water systems. The results
presented here describe the effects that organic solvents have on the
growth of these organisms. The maximal growth rate of Pseudomonas
putida S12, 0.8 h Many organic solvents are toxic to
living organisms because of their devastating effects on biological
membranes (31). This toxicity correlates with the
hydrophobic character of the solvent, expressed by the logarithm of its
partition coefficient between octanol and water (log
PO/W value). Solvents with a log
PO/W value between 1 and 5, like toluene, are
highly toxic to whole cells (30). Due to these toxic
effects, the choice of solvents for whole-cell biotransformations in
two-phase solvent-water systems is limited. Only less-toxic solvents
with higher hydrophobicities (28) can be applied. In the
last decade, however, more and more bacterial strains that can adapt to
toxic organic solvents have been isolated and characterized (5,
15, 26, 34). These solvent-tolerant strains presumably will
become a useful key in the performance of whole-cell biotransformations
in the presence of these toxic, more polar solvents.
In recent years, many efforts have been made to uncover the mechanisms
behind the solvent tolerance of these strains belonging to the genus
Pseudomonas. Up to now different adaptation mechanisms have
been found. Alterations at the level of the cell envelope structure,
which suppress the effects of the solvents on the membrane stability or
limit their rate of diffusion into the cell, have been described
(12, 23, 27, 35). Furthermore, enhanced rates of
phospholipid biosynthesis, speeding up repairing processes, have been
reported (24). Last but not least, active export systems have been shown to exclude the solvent toluene from the cell (6, 16, 19, 27). The active efflux of solvents is an energy-dependent process. Therefore, it should increase the maintenance requirement of
the cells in the presence of solvents. To what extent organic solvents
enhance the energy requirement of the solvent-tolerant strains must
still be determined.
However, although the adaptation mechanisms of some solvent-tolerant
Pseudomonas strains have been studied in detail, no studies have been made of the effects of solvents on growth yields and maintenance requirements of these organisms. We now have determined the
effects of toluene and other solvents on the growth parameters of
Pseudomonas putida S12.
Microorganisms and media.
P. putida S12 was isolated
as a styrene-degrading organism (10). This strain grows in
the presence of a second phase of various organic solvents, even if
these solvents, like toluene, cannot be metabolized (34).
P. putida JK1 is a solvent-sensitive transposon mutant of
P. putida S12. This mutant has inactive sprABC genes which code for the energy-dependent solvent efflux system (19). Both strains were cultivated in a minimal medium as
described by Hartmans et al. (9) with 1.8 g of glucose
per liter as the sole source of carbon and energy. For cultivation on a
solidified medium 3.5 g of yeast extract per liter and 15 g
of agar per liter were added. For the mutant strain 50 mg of kanamycin
per liter was always added to the growth medium.
Batch cultivation.
Batch culture cells were grown in 25-ml
shaken cultures in 250-ml bottles sealed with Mininert valves (Phase
Separations, Waddinxveen, The Netherlands) to prevent evaporation of
the solvent. These valves possess movable Teflon-rubber septa for both
sealing with Teflon and sampling. Different concentrations of toluene were added to the medium and equilibrated at 30°C for at least 12 h. The amount of solvent necessary to achieve a certain amount of solvent in the medium was calculated as follows:
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Copyright © 1999, American Society for Microbiology. All rights reserved.
Effect of Organic Solvents on the Yield of
Solvent-Tolerant Pseudomonas putida S12
ABSTRACT
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Abstract
Introduction
Materials and Methods
Results
Discussion
References
1, was not affected by toluene in
batch cultures, but in chemostat cultures the solvent decreased the
maximal growth rate by nearly 50%. Toluene, ethylbenzene,
propylbenzene, xylene, hexane, and cyclohexane reduced the biomass
yield, and this effect depended on the concentration of the solvent in
the bacterial membrane and not on its chemical structure. The dose
response to solvents in terms of yield was linear up to an
approximately 200 mM concentration of solvent in the bacterial
membrane, both in the wild type and in a mutant lacking an active
efflux system for toluene. Above this critical concentration the yield
of the wild type remained constant at 0.2 g of protein/g of
glucose with increasing concentrations of toluene. The reduction of the
yield in the presence of solvents is due to a maintenance higher by a
factor of three or four as well as to a decrease of the maximum growth
yield by 33%. Therefore, energy-consuming adaptation processes as well
as the uncoupling effect of the solvents reduce the yield of the
tolerant cells.
INTRODUCTION
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
MATERIALS AND METHODS
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
where Vsolvent,addition is the volume of
solvent (in microliters) necessary to achieve the amount of solvent
(msolvent,medium [in milligrams]) in the
medium,
solvent is the density of solvent (in grams per
milliliter), 
water/air is the partition coefficient of
solvent between water and air as given by Amoore and Hautala (2), Vmedium is the volume of the
medium, and Vair is the volume of the air.
Additionally, the concentration of a solvent was controlled by gas
chromatography analysis of the headspace.
1) in a medium saturated with toluene. After
reaching the steady state these cells were used as the inoculum (1%
[vol/vol]) for the batch cultures.
Cultivation of these batch cultures took place in a horizontally
shaking water bath at 30°C. During growth we monitored the optical
density at 560 nm, the CO2 and protein production, and the
consumption of glucose. The maximal errors encountered in the
determination of protein and other parameters were 20 and 10%, respectively.
Continuous cultivation.
Continuous-culture experiments were
performed in a chemostat with a 0.5-liter working volume at 30°C, pH
7.0, 600 rpm, and the dilution rates mentioned. The amount of oxygen in
the culture broth was determined with a Clark-type oxygen electrode.
Various solvents at different concentrations were supplied to the
chemostat via the gas phase by passing a part of the airflow through a
column filled with the solvent (at least 15 cm). The total airflow was kept constantly at 400 ml min1. Headspace samples of the
air entering and leaving the chemostat were analyzed. The solvent
concentrations in the medium were calculated from the concentration in
the headspace by using the partition coefficients given by Amoore and
Hautala (2). To achieve the adaptation towards a certain
solvent concentration the continuous culture was run first at a
dilution rate of 0.05 h1 for at least 12 h and then
switched to the dilution rate of interest. The steady state was reached
after five further exchanges of the volume. From the steady state we
determined the concentration of protein in quintuplicate and the
concentration of glucose remaining in the medium in duplicate. In this
continuous approach the errors of the protein and glucose
determinations were less than 15 and 5%, respectively.
Analytical methods. The amount of the metabolized carbon source, glucose, was determined by high-pressure liquid chromatography analysis of the culture supernatant. The supernatant was filtered via a 0.2-µm-pore-size filter prior to high-pressure liquid chromatography analysis performed at 70°C on an ION-300 column (LC-Service, Emmen, The Netherlands) with 5 mM H2SO4 as an eluent and with refractive index detection (7). Headspace analysis of organic solvents was performed by analyzing 100 µl of the gas phase on a model 437A gas chromatograph (Packard, Delft, The Netherlands) with a 10% SE-30 Chromosorb WHP 80-100 mesh column (Chrompack, Middelburg, The Netherlands). The CO2 concentration was measured via headspace analysis in a model 427 gas chromatograph (Packard) with a Hayesep Q column (Chrompack). Dry weights were determined by drying washed cell suspensions at 105°C for 24 h prior to weighing. The method of Lowry et al. (22) was used to determine the protein concentration with bovine serum albumin as the standard.
Determination of yield and maintenance. We determined the yield by measuring the amount of protein produced per amount of glucose consumed. As protein constitutes 60% of the total cell dry weight in P. putida S12 this value correlates with the amount of biomass produced per amount of glucose consumed.
The maximum growth yields and the maintenance coefficients were determined according to the equation of Pirt (25) from the data determined in the carbon-limited continuous culture as follows: 1/Yobs = (1/Ymax) + (m/µobs). Yobs is the observed growth yield, Ymax is the maximum growth yield, µobs is the observed specific growth rate as set by the dilution rate, and m is the maintenance metabolism rate.Determination of solvent concentrations in the membrane.
The
amount of a solvent accumulating in the bacterial membrane was
calculated from its concentration in the water phase and its log
PO/W, the logarithm of the partition coefficient
of the solvent between octanol and water. For this calibration we made use of the equilibration found by Sikkema et al. (30). This equilibration correlates the log PO/W with the
log PM/B, the logarithm of the partition
coefficient of the solvent between the membrane and the buffer, as
follows: log PM/B = 0.97 × log
PO/W 
0.64. The values for the log
PO/W were obtained from the list reported by
Laane et al. (21).
Chemicals. Toluene, benzene, ethylbenzene, propylbenzene, xylene, hexane, cyclohexane, and hexadecane were obtained from Janssen Chimia (Tilburg, The Netherlands). All other chemicals were commercially available and used without any further purification.
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RESULTS |
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Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
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Growth kinetics of P. putida S12 in batch cultures.
P. putida S12 was precultivated in a continuous culture in a
medium saturated with toluene. These adapted cells were used as the
inoculum for batch cultures, and their growth was monitored. With this
approach cells are already adapted in the beginning of a batch
experiment. Therefore, the response of these cells to a new exposure to
toluene demonstrates the effect of this solvent alone and not the
induction of adaptation mechanisms. The batch growth rate was 0.8 h1 and was not affected by the presence of toluene.
However, variations in the lag phase occurred depending on the
concentration of toluene present. Furthermore, the presence of toluene
led to a lower yield. In the absence and presence of 6.2 mM toluene the
yields observed at the end of the exponential phase were 0.34 and
0.20 g of protein/g of glucose, respectively. Growth kinetics are
presented for cultures growing in either the absence or the presence of
6.2 mM toluene (Fig. 1). The yield and
lag phase obtained for all concentrations of toluene tested are also
shown (see Fig. 4).
|
Effect of toluene on growth of P. putida S12 in
carbon-limited continuous culture.
P. putida S12 was
cultivated in a carbon-limited chemostat at different dilution rates in
the presence and absence of 6.2 mM toluene. After the cells had reached
the steady state we determined the protein content and the
concentration of glucose as the growth-limiting substrate (Fig.
2). In the absence of toluene the washout
occurred at a dilution rate above 0.72 h1. In the
presence of 6.2 mM toluene, however, this washout occurred at a lower
growth rate, and it was not possible to obtain a steady state at
dilution rates above 0.4 h1. The presence of toluene also
led to smaller amounts of biomass at all dilution rates. It was not
possible to detect protein in the culture supernatant at all dilution
rates tested.
|
1, respectively, and the
maximum growth yields were 0.33 and 0.22 g of protein/g of
glucose, respectively.
|
) and presence (
) of 6.2 mM toluene. The linear regression values (determined before washout
occurs) are as follows: 1/Y = (0.023/D) + 3.00 (in the absence of toluene) and 1/Y = (0.076/D) + 4.57 (in the presence of toluene).
1 in the presence of various
concentrations of toluene. Figure 4 shows
the effect of toluene on the yield. Up to a 3 mM concentration of
toluene in the medium, the yield decreases linearly with increasing concentrations of toluene. Above this concentration of toluene, the
yield remained nearly constant at 0.21 g of protein/g of glucose. This dose response is similar to results obtained in batch cultures.
|
) is
taken as the period of time from the inoculation of batch cultures
until an increase in the optical density was observed. Yields were
determined in batch cultures at the end of the exponential phase (
)
and from cells growing in a glucose-limited chemostat at a dilution
rate of 0.2 hEffects of different solvents on the yield of P. putida
S12.
P. putida S12 was grown in continuous cultures at a
dilution rate of 0.2 h1. Various aromatic and aliphatic
organic solvents were added at different concentrations. We determined
the concentration of the solvent in the medium and calculated the
corresponding concentration in the bacterial membrane as described in
Materials and Methods. We plotted the yield of P. putida S12
against these membrane concentrations (Fig.
5). The plot shows a direct correlation
between the yields observed and the concentrations of solvents in the
membrane, irrespective of the solvent tested.
|
), hexane (+), cyclohexane
(
), and hexadecane (
) were added at different concentrations. The
theoretical concentrations of the solvents in the bacterial membrane
were calculated (see Materials and Methods), and the yields were
plotted against these concentrations.
Effect of toluene on the yield of the solvent-sensitive mutant P. putida JK1. The effects of solvents were also studied in the solvent-sensitive mutant P. putida JK1. This mutant is lacking the operon for the solvent efflux system. The cells were grown in batch cultures in the presence of various concentrations of toluene. The cells were transferred from lower toluene concentrations to higher ones in small concentration steps of about 0.5 mM each at the end of the exponential phase. In this way the growth of the mutant could be obtained at toluene concentrations up to 3.2 mM. Above this concentration no growth of the mutant strain was observed, while the wild-type strain tolerated a 6.2 mM concentration of toluene. For both strains we monitored the protein production and glucose consumption in the presence of various concentrations of toluene. From these data, yields were calculated and plotted against the concentration of toluene (Fig. 6). Up to a 2 mM concentration of toluene the yields of both strains were similar. Higher concentrations reduced the yield of the mutant to zero.
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DISCUSSION |
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Abstract
Introduction
Materials and Methods
Results
Discussion
References
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The growth-inhibiting effect of organic solvents on microorganisms has been reported repeatedly (28, 31). The presence of solvents may lead to a reduction in the maximum growth rate, but cells which can adapt to solvents can achieve the same maximum growth rate in the presence of solvents (3, 11, 15, 26). These results were obtained by employing batch systems, and we also observed that in batch cultivation no effect of toluene on the growth rate occurred. However, these results in the case of glucose as a carbon source may be misleading, as it was reported previously that gluconic acid transiently accumulates during the cultivation of P. putida on glucose (8). Under more-defined conditions in continuous cultures we observed that the maximal growth rate is strongly affected by the presence of solvents. This reduction was not caused by oxygen limitation, as we had determined that sufficient oxygen was present in all cases. We speculate that the reduction of the maximum growth rate in the presence of toluene is caused by a reduction in the affinity of the cells for glucose. Such a change in the affinity for glucose may be caused by the mechanisms of adaptation to toluene that reduce the permeability and change the structure of the cell envelope (18, 23, 27, 36).
The effects of solvents on the biomass yield have been implied in previous reports (1, 3, 27). From these reports it can be deduced that the presence of toluene leads to reduced yields. We observed similar results. In our batch cultures the yield as affected by toluene dropped by 30%.
In continuous cultures, yields and maintenance coefficients were obtained for P. putida S12 grown under glucose limitation in either the presence or the absence of toluene and other solvents. In the absence of solvents, both the maximal yield and the maintenance coefficient were similar to those reported for other Pseudomonas species growing on glucose under aerobic conditions (8, 25, 32, 33).
The presence of toluene decreased the yield and increased the maintenance coefficient. These effects will be caused both by energy-consuming adaptation mechanisms and by a less effective energy metabolism in the presence of solvents. Specific energy-consuming processes include the active export of solvents in P. putida S12 (16, 19) and possibly an enhanced phospholipid biosynthesis rate as observed in P. putida Idaho (24). Ineffective energy metabolism will occur due to the uncoupling character of organic solvents (4, 13, 29, 30) and by disturbing effects of the solvents on the energy-transducing proteins (29, 30) as observed in nontolerant microorganisms.
The dose-response effect of toluene on the overall yield of P. putida S12 was studied in both batch and chemostat cultures. The yield decreased linearly with increasing concentrations (up to 3 mM) of toluene. Above this concentration, no further drop in the yield occurred. The adaptation of P. putida S12 to toluene has been observed at 3 mM or higher concentrations of toluene. Adaptation mechanisms triggered at this concentration include changes in the fatty acid profiles of membranes (35) and the active efflux of solvents from the membrane (20). Cells precultured at this concentration or higher ones not only survived the presence of a second phase of toluene (34), but they also showed an enhanced resistance towards various antibiotics (17). Consequently, a toluene concentration of 3 mM is critical. Below 3 mM cells do not react to toluene and thus are slightly affected by the solvent. Above this concentration of toluene, various mechanisms come into operation to protect cells from excessive damage.
The constant yield values observed at toluene concentrations of 3 to 6 mM may indicate either that (i) the energy requirement of the adaptation mechanisms acting at these concentrations is very limited or (ii) the systems require substantial energy input but are compensated by the effective removal of toluene from the cell.
The results obtained for the wild type were confirmed in experiments with the solvent-sensitive mutant P. putida JK1. At low concentrations of toluene, the effects of solvents on the yield were similar in the wild type and the solvent-sensitive mutant P. putida JK1. The slightly lower yield of the mutant strain is caused by the presence of kanamycin as the selective marker. The mutant lacks the energy-consuming solvent efflux system. Therefore, the reduction of the yield observed at low concentrations of toluene cannot be caused by the energy requirement of the active efflux system.
Our results on yields of P. putida S12 as affected by various other solvents show that the dose-response effect of solvents is the same when the actual concentration in the bacterial membrane is taken as the dose. Therefore, not the chemical structure but the amount of solvent accumulated in the bacterial membrane determines the effect of a solvent on the yield. Hence, the results found for toluene can be used for other solvents as well. It has been reported earlier that the concentration of solvents in bacterial membranes correlates directly with changes in the fatty acid profile and with the reduction of the maximal growth rate (14). In artificial membranes the same concentration of a solvent in the membrane results in the same expansion of the membrane. This membrane concentration also determines the release of ions and the effect on the proton and electrical gradients (30).
We conclude that, at low concentrations, the effects of solvents on P. putida S12 are not different from their effects on any other cell. The difference between solvent-tolerant and -intolerant cells seems to be that the effects of solvents are counterbalanced at higher concentrations by specific mechanisms. We suggest that these mechanisms keep the actual concentrations of solvents in the membrane constant. We think that this constant concentration of solvent in the membrane is approximately 200 mM, the concentration reached when adaptation starts.
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ACKNOWLEDGMENTS |
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This work was financially supported in the framework of an industrially relevant research program of the Association of Biotechnology Centers in The Netherlands.
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FOOTNOTES |
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* Corresponding author. Mailing address: Division of Industrial Microbiology, Department of Food Technology and Nutritional Sciences, Wageningen Agricultural University, P.O. Box 8129, 6700 EV Wageningen, The Netherlands. Phone: 31 317 484412. Fax: 31 317 484978. E-mail: Sonja.Isken{at}imb.ftns.wau.nl.
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REFERENCES |
|---|
|
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Abstract
Introduction
Materials and Methods
Results
Discussion
References
|
|---|
| 1. | Abe, A., A. Inoue, R. Usami, K. Moriya, and K. Horikoshi. 1995. Degradation of polyaromatic hydrocarbons by organic solvent-tolerant bacteria from deep sea. Biosci. Biotechnol. Biochem. 59:1154-1156[Medline]. |
| 2. | Amoore, J. E., and E. Hautala. 1983. Odor as an aid to chemical safety: odor thresholds compared with threshold limit values and volatilities for 214 industrial chemicals in air and water dilution. J. Appl. Toxicol. 3:272-290[Medline]. |
| 3. | Aono, R., M. Ito, A. Inoue, and K. Horikoshi. 1992. Isolation of novel toluene-tolerant strain Pseudomonas aeruginosa. Biosci. Biotechnol. Biochem. 56:145-146. |
| 4. | Cartwright, C. P., J.-R. Juroszek, M. J. Beavan, F. M. S. Ruby, S. M. F. de Morais, and A. H. Rose. 1986. Ethanol dissipates the proton motive force across the plasma membrane of Saccharomyces cerevisiae. J. Gen. Microbiol. 132:369-377. |
| 5. |
Cruden, D. L.,
J. H. Wolfram,
R. D. Rogers, and D. T. Gibson.
1992.
Physiological properties of a Pseudomonas strain which grows with p-xylene in a two-phase (organic-aqueous) medium.
Appl. Environ. Microbiol.
58:2723-2729 |
| 6. | Fukumori, F., H. Hirayama, H. Takami, A. Inoue, and K. Horikoshi. 1998. Isolation and transposon mutagenesis of a Pseudomonas putida KT2442 toluene-resistant variant: involvement of an efflux system in solvent resistance. Extremophiles 2:395-400. [Medline] |
| 7. | Grobben, G. J., J. Sikkema, M. R. Smith, and J. A. M. de Bont. 1995. Production of extracellular polysaccharides by Lactobacillus delbrueckii ssp. bulgaricus NCFB 2772 grown in a chemically defined medium. J. Appl. Bacteriol. 79:103-107. |
| 8. | Hack, C. J., J. M. Woodley, M. D. Lilly, and J. M. Liddell. 1994. The production of Pseudomonas putida for the hydroxylation of toluene to its cis-glycol. Appl. Microbiol. Biotechnol. 41:495-499. |
| 9. |
Hartmans, S.,
J. P. Smits,
M. J. van der Werf,
F. Volkering, and J. A. M. de Bont.
1989.
Metabolism of styrene oxide and 2-phenylethanol in the styrene-degrading Xanthobacter strain 124X.
Appl. Environ. Microbiol.
55:2850-2855 |
| 10. |
Hartmans, S.,
M. J. van der Werf, and J. A. M. de Bont.
1990.
Bacterial degradation of styrene involving a novel flavin adenine dinucleotide-dependent styrene monooxygenase.
Appl. Environ. Microbiol.
56:1347-1351 |
| 11. |
Heipieper, H. J., and J. A. M. de Bont.
1994.
Adaptation of Pseudomonas putida S12 to ethanol and toluene at the level of fatty acid composition of membranes.
Appl. Environ. Microbiol.
60:4440-4444 |
| 12. |
Heipieper, H.-J.,
R. Diefenbach, and H. Keweloh.
1992.
Conversion of cis unsaturated fatty acids to trans, a possible mechanism for the protection of phenol-degrading Pseudomonas putida P8 from substrate toxicity.
Appl. Environ. Microbiol.
58:1847-1852 |
| 13. |
Heipieper, H.-J.,
H. Keweloh, and H.-J. Rehm.
1991.
Influence of phenols on growth and membrane permeability of free and immobilized Escherichia coli.
Appl. Environ. Microbiol.
57:1213-1217 |
| 14. | Heipieper, H. J., B. Loffeld, H. Keweloh, and J. A. M. de Bont. 1995. The cis/trans isomerization of unsaturated fatty acids in Pseudomonas putida S12: an indicator for environmental stress due to organic compounds. Chemosphere 30:1041-1051. |
| 15. | Inoue, A., and K. Horikoshi. 1989. A Pseudomonas thrives in high concentrations of toluene. Nature 338:264-266. |
| 16. | Isken, S., and J. A. M. de Bont. 1996. Active efflux of toluene in a solvent-resistant bacterium. J. Bacteriol. 90:6056-6058. |
| 17. | Isken, S., P. M. A. C. Santos, and J. A. M. de Bont. 1997. Effect of solvent adaptation on the antibiotic resistance in Pseudomonas putida S12. Appl. Microbiol. Biotechnol. 48:642-647. |
| 18. | Isken, S., and J. A. M. de Bont. 1998. Solvent-tolerant bacteria. Extremophiles 2:229-238. [Medline] |
| 19. |
Kieboom, J.,
J. J. Dennis,
J. A. M. de Bont, and G. J. Zylstra.
1998.
Identification and molecular characterization of an efflux pump involved in Pseudomonas putida S12 solvent tolerance.
J. Biol. Chem.
273:85-91 |
| 20. |
Kieboom, J.,
J. J. Dennis,
J. A. M. de Bont, and G. J. Zylstra.
1998.
Active efflux of organic solvents in Pseudomonas putida S12 is induced by solvents.
J. Bacteriol.
180:6769-6772 |
| 21. | Laane, C., S. Boeren, R. Hilhorst, and C. Veeger. 1987. Optimization of biocatalysis in organic media. In C. Laane, J. Tramper, and M. D. Lilly (ed.), Biocatalysis in organic media. Elsevier Science Publishers, Amsterdam, The Netherlands. |
| 22. |
Lowry, O. H.,
N. J. Rosebrough,
A. L. Farr, and R. J. Randall.
1951.
Protein measurement with the Folin phenol reagent.
J. Biol. Chem.
193:265-275 |
| 23. | Pinkart, H. C., J. W. Wolfram, R. Rogers, and D. C. White. 1996. Cell envelope changes in solvent-tolerant and solvent-sensitive Pseudomonas putida strains following exposure to o-xylene. Appl. Environ. Microbiol. 62:1129-1132[Abstract]. |
| 24. |
Pinkart, H. C., and D. C. White.
1997.
Phospholipid biosynthesis and solvent tolerance in Pseudomonas putida strains.
J. Bacteriol.
179:4219-4226 |
| 25. | Pirt, S. J. 1975. Principles of microbe and cell cultivation, p. 63-70. Wiley, New York, N.Y. |
| 26. |
Ramos, J. L.,
E. Duque,
M.-J. Huertas, and A. Haïdour.
1995.
Isolation and expansion of the catabolic potential of a Pseudomonas putida strain able to grow in the presence of high concentrations of aromatic hydrocarbons.
J. Bacteriol.
177:3911-3916 |
| 27. |
Ramos, J. L.,
E. Duque,
J. J. Rodriguez-Herva,
P. Godoy,
A. Haïdour,
F. Reyes, and A. Fernandez-Barrero.
1997.
Mechanisms for solvent tolerance in bacteria.
J. Biol. Chem.
272:3887-3890 |
| 28. | Salter, G. J., and D. B. Kell. 1995. Solvent selection for whole cell biotransformation in organic media. Crit. Rev. Biotechnol. 15:139-177[Medline]. |
| 29. |
Sikkema, J.,
B. Poolman,
W. N. Konings, and J. A. M. de Bont.
1992.
Effects of the membrane action of tetraline on the function and structural properties of artificial and bacterial membranes.
J. Bacteriol.
174:2986-2992 |
| 30. |
Sikkema, J.,
J. A. M. de Bont, and B. Poolman.
1994.
Interactions of cyclic hydrocarbons with biological membranes.
J. Biol. Chem.
269:8022-8028 |
| 31. |
Sikkema, J.,
J. A. M. de Bont, and B. Poolman.
1995.
Mechanisms of membrane toxicity of hydrocarbons.
Microbiol. Rev.
59:201-222 |
| 32. |
Verdoni, N.,
M. A. Aon, and J. M. Lebeault.
1992.
Metabolic and energetic control of Pseudomonas mendocina growth during transitions from aerobic to oxygen-limited conditions in chemostat cultures.
Appl. Environ. Microbiol.
58:3150-3156 |
| 33. | Wang, K.-W., B. C. Baltzis, and G. A. Lewandowski. 1996. Kinetics of phenol biodegradation in the presence of glucose. Biotechnol. Bioeng. 51:87-94. |
| 34. |
Weber, F. J.,
L. P. Ooijkaas,
R. M. W. Schemen,
S. Hartmans, and J. A. M. de Bont.
1993.
Adaptation of Pseudomonas putida S12 to high concentrations of styrene and other organic solvents.
Appl. Environ. Microbiol.
59:3502-3504 |
| 35. |
Weber, F. J.,
S. Isken, and J. A. M. de Bont.
1994.
Cis/trans isomerization of fatty acids as a defense mechanism of Pseudomonas putida strains to toxic concentrations of toluene.
Microbiology
140:2013-2017 |
| 36. | Weber, F. J., and J. A. M. de Bont. 1996. Adaptation mechanisms of microorganisms to the toxic effects of organic solvents on membranes. Biochim. Biophys. Acta 1286:225-245[Medline]. |
Applied and Environmental Microbiology, June 1999, p. 2631-2635, Vol. 65, No. 6
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