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Isolation and Characterization of rpoS from a Pathogenic Bacterium, Vibrio vulnificus: Role of  S
in Survival of Exponential-Phase Cells under Oxidative Stress.Kyung-Je Park, 2004.A gene homologous to rpoS was cloned from a fatal human pathogen, Vibrio vulnificus . The functional role of rpoS in V . vulnificus was accessed by using an rpoS knockout mutant strain . This mutant was impaired in terms of the ability to survive under oxidative stress, nutrient starvation, UV irradiation, or acidic conditions . The increased susceptibility of the V . vulnificus mutant in the exponential phase to H2O2 was attributed to the reduced activity of hydroperoxidase I (HPI) . Although S
synthesis was induced and HPI activity reached the maximal level in
the stationary phase, the mutant in the stationary phase showed the
same susceptibility to H2O2 as the wild-type
strain in the stationary phase . In addition, HPII activity, which is
known to be controlled by
S
in Escherichia coli, was not detectable in V . vulnificus strains
under the conditions tested . The mutant in the exponential phase
complemented with multiple copies of either the rpoS or katG
gene of V . vulnificus recovered both resistance to H2O2
and HPI activity compared with the control strain . Expression of
the katG gene encoding HPI in V . vulnificus was monitored by
using a katG::luxAB transcriptional fusion . The expression
of this gene was significantly reduced by deletion of
S
in both the early exponential and late stationary phases . Thus,
S
is necessary for increased synthesis and activity of HPI, and
S
is required for exponentially growing V . vulnificus to develop
the ability to survive in the presence of H2O2 .
Genetic Typing of Shiga Toxin 2 Variants of Escherichia coli by PCR-Restriction Fragment Length Polymorphism Analysis. Liesbet De Baets, 2004. Genetic and Biochemical Characterization of the F-ATPase Operon from Streptococcus sanguis 10904. Wendi L. Kuhnert, 2003.Oral streptococci utilize an F-ATPase to regulate cytoplasmic pH . Previous studies have shown that this enzyme is a principal determinant of aciduricity in the oral streptococcal species Streptococcus sanguis and Streptococcus mutans . Differences in the pH optima of the respective ATPases appears to be the main reason that S . mutans is more tolerant of low pH values than S . sanguis and hence pathogenic . We have recently reported the genetic arrangement for the S . mutans operon . For purposes of comparative structural biology we have also investigated the F-ATPase from S . sanguis . Here, we report the genetic characterization and expression in Escherichia coli of the S . sanguis ATPase operon . Sequence analysis showed a gene order of atpEBFHAGDC and that a large intergenic space existed upstream of the structural genes . Activity data demonstrate that ATPase activity is induced under acidic conditions in both S . sanguis and S . mutans; however, it is not induced to the same extent in the nonpathogenic S . sanguis . Expression studies with an atpD deletion strain of E . coli showed that S . sanguis-E . coli hybrid enzymes were able to degrade ATP but were not sufficiently functional to permit growth on succinate minimal media . Hybrid enzymes were found to be relatively insensitive to inhibition by dicyclohexylcarbodiimide, indicating loss of productive coupling between the membrane and catalytic subunits . Anaerobic Degradation of Ethylbenzene by a New Type of Marine Sulfate-Reducing Bacterium. Olaf Kniemeyer, 2003.Anaerobic degradation of the aromatic hydrocarbon ethylbenzene was studied with sulfate as the electron acceptor . Enrichment cultures prepared with marine sediment samples from different locations showed ethylbenzene-dependent reduction of sulfate to sulfide and always contained a characteristic cell type that formed gas vesicles towards the end of growth . A pure culture of this cell type, strain EbS7, was isolated from sediment from Guaymas Basin (Gulf of California) . Complete mineralization of ethylbenzene coupled to sulfate reduction was demonstrated in growth experiments with strain EbS7 . Sequence analysis of the 16S rRNA gene revealed a close relationship between strain EbS7 and the previously described marine sulfate-reducing strains NaphS2 and mXyS1 (similarity values, 97.6 and 96.2%, respectively), which grow anaerobically with naphthalene and m-xylene, respectively . However, strain EbS7 did not oxidize naphthalene, m-xylene, or toluene . Other compounds utilized by strain EbS7 were phenylacetate, 3-phenylpropionate, formate, n-hexanoate, lactate, and pyruvate . 1-Phenylethanol and acetophenone, the characteristic intermediates in anaerobic ethylbenzene degradation by denitrifying bacteria, neither served as growth substrates nor were detectable as metabolites by gas chromatography-mass spectrometry in ethylbenzene-grown cultures of strain EbS7 . Rather, (1-phenylethyl)succinate and 4-phenylpentanoate were detected as specific metabolites in such cultures . Formation of these intermediates can be explained by a reaction sequence involving addition of the benzyl carbon atom of ethylbenzene to fumarate, carbon skeleton rearrangement of the succinate moiety (as a thioester), and loss of one carboxyl group . Such reactions are analogous to those suggested for anaerobic n-alkane degradation and thus differ from the initial reactions in anaerobic ethylbenzene degradation by denitrifying bacteria which employ dehydrogenations .
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