Isolation and characterization of Vitreoscilla mutant defective in catalase-peroxidase hydropreroxidase 1
Isolation and characterization of Vitreoscilla mutant defective in catalase-peroxidase hydroperoxidase 1
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Mutants of an obligate aerobic bacterium, Vitreoscilla, that have deficiency in heatlabile catalaseperoxidase hydroperoxidase I(HPI) were created by EMS treatment. The catalaseperoxidase HPIdeficient mutant showed substantially lower peroxidase activity in exponential and midstationary phase compared with the wild type strain. In late stationary phase, the mutant exhibited no peroxidase activity. Peroxidase deficiency in the mutant was revealed by polyacrylamide gels stained for peroxidase activity. Characteristically, catalase levels in the mutant increased about 14and 8fold during growth in the exponential and stationary phases, respectively, compared to those in the wild type, suggesting a compensatory effect for protection from H2O2 toxicity. The mutant showed differences in physiology from the wild type: retardation in growth rate and decrease in oxygen consumption. Both the wild type and the catalaseperoxidase HPIdeficient mutant of Vitreoscilla had lower growth rates in media containing increasing H2O2 concentrations. However, the mutant exhibited an additionally decreased growth rate after 6 to 8 h of growth compared to the wild type. The wild type was resistent up to 20 mM H2O2, whereas the mutant was very sensitive to high concentrations of exogenous H2O2. Although elevated catalase levels would provide protection of the bacteria from the deleterious effect of H2O2, it did not appear to be complete. Cellfree extracts of the mutant showed decreased NADH oxidation *Corresponding authorPhone : +82-63-270-3361, Fax +82-63-270-3362 E-mail : park253@chonbuk.ac.krrates and higher accumulation of H2O2 during this oxidation. These results may account for the impaired growth and earlier onset of death phase by the catalaseperoxidase HPIdeficient mutant of Vitreoscilla.
Abstract
Mutants of an obligate aerobic bacterium, Vitreoscilla, that have deficiency in heatlabile catalaseperoxidase hydroperoxidase I(HPI) were created by EMS treatment. The catalaseperoxidase HPIdeficient mutant showed substantially lower peroxidase activity in exponential and midstationary phase compared with the wild type strain. In late stationary phase, the mutant exhibited no peroxidase activity. Peroxidase deficiency in the mutant was revealed by polyacrylamide gels stained for peroxidase activity. Characteristically, catalase levels in the mutant increased about 14and 8fold during growth in the exponential and stationary phases, respectively, compared to those in the wild type, suggesting a compensatory effect for protection from H2O2 toxicity. The mutant showed differences in physiology from the wild type: retardation in growth rate and decrease in oxygen consumption. Both the wild type and the catalaseperoxidase HPIdeficient mutant of Vitreoscilla had lower growth rates in media containing increasing H2O2 concentrations. However, the mutant exhibited an additionally decreased growth rate after 6 to 8 h of growth compared to the wild type. The wild type was resistent up to 20 mM H2O2, whereas the mutant was very sensitive to high concentrations of exogenous H2O2. Although elevated catalase levels would provide protection of the bacteria from the deleterious effect of H2O2, it did not appear to be complete. Cellfree extracts of the mutant showed decreased NADH oxidation *Corresponding authorPhone : +82-63-270-3361, Fax +82-63-270-3362 E-mail : park253@chonbuk.ac.krrates and higher accumulation of H2O2 during this oxidation. These results may account for the impaired growth and earlier onset of death phase by the catalaseperoxidase HPIdeficient mutant of Vitreoscilla.
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