Journal of Microbiology

Journal Article

The inability of Bacillus licheniformis perR mutant to grow is mainly due to the lack of PerR-mediated fur repression: Jung-Hoon Kim , Yoon-Mo Yang , Chang-Jun Ji , Su-Hyun Ryu , Young-Bin Won , Shin-Yeong Ju , Yumi Kwon , Yeh-Eun Lee , Hwan Youn , Jin-Won Lee; J. Microbiol. 2017;55(6):457-463. Published online April 22, 2017; DOI: https://doi.org/10.1007/s12275-017-7051-x

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Abstract: PerR, a member of Fur family protein, is a metal-dependent H2O2 sensing transcription factor that regulates genes in-volved in peroxide stress response. Industrially important bac-terium Bacillus licheniformis contains three PerR-like pro-teins (PerRBL, PerR2, and PerR3) compared to its close rela-tive Bacillus subtilis. Interestingly, unlike other bacteria in-cluding B. subtilis, no authentic perRBL null mutant could be established for B. licheniformis. Thus, we constructed a con-ditional perRBL mutant using a xylose-inducible promoter, and investigated the genes under the control of PerRBL. PerRBL regulon genes include katA, mrgA, ahpC, pfeT, hemA, fur, and perR as observed for PerRBS. However, there is some variation in the expression levels of fur and hemA genes be-tween B. subtilis and B. licheniformis in the derepressed state. Furthermore, katA, mrgA, and ahpC are strongly induced, whereas the others are only weakly or not induced by H2O2 treatment. In contrast to the B. subtilis perR null mutant which frequently gives rise to large colony phenotype mainly due to the loss of katA, the suppressors of B. licheniformis perR mutant, which can form colonies on LB agar, were all cata-lase-positive. Instead, many of the suppressors showed in-creased levels of siderophore production, suggesting that the suppressor mutation is linked to the fur gene. Consistent with this, perR fur double mutant could grow on LB agar without Fe supplementation, whereas perR katA double mutant could only grow on LB agar with Fe supplementation. Taken toge-ther, our data suggest that in B. licheniformis, despite the si-milarity in PerRBL and PerRBS regulon genes, perR is an essen-tial gene required for growth and that the inability of perR null mutant to grow is mainly due to elevated expression of Fur.

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Pseudomonas aeruginosa MdaB and WrbA are Water-soluble Two-electron Quinone Oxidoreductases with the Potential to Defend against Oxidative Stress: Laura K Green , Anne C La Flamme , David F Ackerley; J. Microbiol. 2014;52(9):771-777. Published online August 2, 2014; DOI: https://doi.org/10.1007/s12275-014-4208-8

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Abstract: Water-soluble quinone oxidoreductases capable of reducing quinone substrates via a concerted two-electron mechanism have been implicated in bacterial antioxidant defence. Twoelectron transfer avoids formation of dangerously reactive semi-quinone intermediates, moreover previous work in Pseudomonas putida indicated a direct protective effect for the quinols generated by an over-expressed oxidoreductase. Here, the Pseudomonas aeruginosa orthologs of five quinone oxidoreductases – MdaB, ChrR, WrbA, NfsB, and NQO1 – were tested for their possible role in defending P. aeruginosa against H2O2 challenge. In in vitro assays, each enzyme was shown to reduce quinone substrates with only minimal semiquinone formation. However, when each was individually over-expressed in P. aeruginosa no overt H2O2-protective phenotype was observed. It was shown that this was due to a masking effect of the P. aeruginosa catalase, KatA; in a katA mutant, H2O2 challenged strains over-expressing the WrbA and MdaB orthologs grew significantly better than the empty plasmid control. A growth advantage was also observed for H2O2 challenged P. putida strains over-expressing P. aeruginosa wrbA, mdaB or katA. Despite not conferring a growth advantage to wild type P. aeruginosa, it is possible that these quinone oxidoreductases defend against H2O2 toxicity at lower concentrations.

Involvement of Alternative Oxidase in the Regulation of Growth, Development, and Resistance to Oxidative Stress of Sclerotinia sclerotiorum: Ting Xu , Fei Yao , Wu-Sheng Liang , Yong-Hong Li , Dian-Rong Li , Hao Wang , Zheng-Yi Wang; J. Microbiol. 2012;50(4):594-602. Published online August 25, 2012; DOI: https://doi.org/10.1007/s12275-012-2015-7

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Abstract: Sclerotinia sclerotiorum is a cosmopolitan, filamentous, fungal pathogen that can cause serious disease in many kinds of crops. Alternative oxidase is the terminal oxidase of the alternative mitochondrial respiratory pathway in fungi and higher plants. We report the presence of this alternative pathway respiration and demonstrate its expression in two isolates of S. sclerotiorum under unstressed, normal culture conditions. Application of salicylhydroxamic acid, a specific inhibitor of alternative oxidase, severely inhibited the mycelial growth of S. sclerotiorum both on potato dextrose agar plates and in liquid culture media. Inhibition of alternative oxidase could influence the growth pattern of S. sclerotiorum, as salicylhydroxamic acid treatment induced obvious aerial mycelia growing on potato dextrose agar plates. Under the treatment with salicylhydroxamic acid, S. sclerotiorum formed sclerotia much more slowly than the control. Treatment with hydrogen peroxide in millimolar concentrations greatly decreased the growth rate of mycelia and delayed the formation of sclerotia in both tested S. sclerotiorum isolates. As well, this treatment obviously increased their alternative pathway respiration and the levels of both mRNA and protein of the alternative oxidase. These results indicate that alternative oxidase is involved in the regulation of growth, development, and resistance to oxidative stress of S. sclerotiorum.

YlaC is an Extracytoplasmic Function (ECF) Sigma Factor Contributing to Hydrogen Peroxide Resistance in Bacillus subtilis: Han-Bong Ryu , Inji Shin , Hyung-Soon Yim , Sa-Ouk Kang; J. Microbiol. 2006;44(2):206-216.; DOI: https://doi.org/2363 [pii]

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Abstract: In this study, we have attempted to characterize the functions of YlaC and YlaD encoded by ylaC and ylaD genes in Bacillus subtilis. The GUS reporter gene, driven by the yla operon promoter, was expressed primarily during the late exponential and early stationary phase, and its expression increased as the result of hydrogen peroxide treatment. Northern and Western blot analyses revealed that the level of ylaC transcripts and YlaC increased as the result of challenge with hydrogen peroxide. A YlaC-overexpressing strain evidenced hydrogen peroxide resistance and a three-fold higher peroxidase activity as compared with a deletion mutant. YlaC-overexpressing and YlaD-disrupted strains evidenced higher sporulation rates than were observed in the YlaC-disrupted and YlaD-overexpressing strains. Analyses of the results of native polyacrylamide gel electrophoresis of recombinant YlaC and YlaD indicated that interaction between YlaC and YlaD was regulated by the redox state of YlaD in vitro. Collectively, the results of this study appear to suggest that YlaC regulated by the YlaD redox state, contribute to oxidative stress resistance in B. subtilis.

Isolation of the Regulator Gene Responsible for Overproduction of Catalase A in H 2 O 2 -resistant Mutant of Streptomyces coelicolor: Ji-Sook Hahn , So-Young Oh , Keith F. Chater , Jung-Hye Roe; J. Microbiol. 2000;38(1):18-23.

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Abstract: Streptomyces coelicolor produces three kinds of catalases to cope with oxidative stress and to allow nor-mal differentiation. Catalase A is the major vegetative catalase which functions in removing hydrogen peroxide generated during the process of aerobic metabolism. To understand the regulatory mechanism of response against oxidative stress, hydrogen peroxide-resistant mutant (HR40) was isolated from S. coelicolor J1501 following UV mutagenesis. The mutant overproduced catalase A more than 50-fold compared with the wild type. The mutation locus catR1 was mapped closed to the mthB2 locus by genetic crossings. An ordered cosmid library of S. coelicolor encompassing the mthB2 locus was used to isolate the regulator gene (catR) which represses catalase overproduction when introduced into HR40. A candidate catR gene was found to encode a Fur-like protein of 138 amino acids (15319 Da).

Monitoring and Characterization of Bacterial Contamination in a High-Purity Water System Used for Semiconductor Manufacturing: In Seop Kim , Geon-Hyoung Lee , Kye Joon Lee; J. Microbiol. 2000;38(2):99-104.

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Abstract: Hydrogen peroxide has been used in cleaning the piping of an advanced high-purity water system that supplies ultra-high purity water (UHPW) for 16 megabyte DRAM semiconductor manufacturing. The level of hydrogen peroxide-resistant bacteria in UHPW water was monitored prior to and after disinfecting the piping with hydrogen peroxide. Most of the bacteria isolated after hydrogen peroxide disinfection were highly resistant to hydrogen peroxide. However, the percentage of resistant bacteria decreased with time. The hydrogen peroxide-resistant bacteria were identified as Micrococcus luteus, Bacillus cereus, Alcaligenes latus, Xanthomonas sp. and Flavobacterium indologenes. The susceptibility of the bacteria to hydrogen peroxide was tested as either planktonic cells or attached cells on glass. Attached bacteria as the biofilm on glass exhibited increased hydrogen peroxide resistance, with the resistance increasing with respect to the age of the biofilm. These results indicate that bacteria resistant to hydrogen peroxide play an important role in biofilm regrowth on piping after hydrogen peroxide treatment. In order to optimize the cleaning strategy for piping of the high-purity water system, the disinfecting effect of hydrogen peroxide and peracetic acid on the bacteria was evaluated. The combined use of hydrogen peroxide and peracetic acid was very effective in killing attached bacteria as well as planktonic bacteria.

Glutathione Content and the Activities of GlutathioneSynthesizing Enzymes in Fission Yeast are Modulated by Oxidative Stress: Yuk-Young Lee , Su-Jung Kim , Eun-Hee Park , Chang-Jin Lim; J. Microbiol. 2003;41(3):248-251.

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Abstract: Glutathione (GSH) is an important factor in determining tolerance against oxidative stress in living organisms. It is synthesized in two sequential reactions catalyzed by [gamma]-glutamylcysteine synthetase (GCS) and glutathione synthetase (GS) in the presence of ATP. In this work, the effects of three different oxidative stresses were examined on GSH content and GSH-related enzyme activities in the fission yeast Schizosaccharomyces pombe. GSH content in S. pombe was significantly enhanced by treatment with hydrogen peroxide, [beta]-naphthoflavone (BNF) and tert-butylhydroquinone (BHQ). Simultaneously, they greatly induced GCS and GS activity. However, they did not have any effects on glutathione reductase activity. These results suggest that GCS and GS activities in S. pombe are upregulated by oxidative stress.

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