Purification, characterization and cloning of a periplasmic catalase from B abortus and the role it plays in the pathogenesis of Brucella

Sha, Zheng yu

doi:10.31274/rtd-180813-12622

Cited by 9 publications

(8 citation statements)

References 83 publications

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“…Recent studies have shown, however, that Brucella grown in macrophages undergo a significant change in protein expression relative to their in vitro grown counterparts (Lin and Ficht, 1995; Rafie‐Kolpin et al ., 1996). Many of these macrophage‐induced proteins are thought to participate in stress response and survival (Latimer et al ., 1992; Tatum et al ., 1992; Sha et al ., 1994; Elzer et al ., 1994; Lin and Ficht, 1995). An intriguing possibility may be that HF‐I participates in the regulation of some or all of these molecules or, alternatively, another global regulatory factor involved in their co‐ordinate expression.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, no essential contributors to macrophage stress adaptation have been identified to date for the brucellae. Moreover, mutations in genes encoding predicted Brucella stress response proteins have little or no effect on pathogenesis, as judged by replication in cultured macrophages or colonization of a murine model of infection (Latimer et al ., 1992; Tatum et al ., 1992; 1993; 1994; Elzer et al ., 1994; 1996; Sha, 1994; Köhler et al ., 1996). Thus, the genetic basis for the capacity of these highly successful intracellular pathogens to maintain prolonged residence in host macrophages is presently unclear.…”

Section: Introductionmentioning

confidence: 99%

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The Brucella abortus host factor I (HF‐I) protein contributes to stress resistance during stationary phase and is a major determinant of virulence in mice

Robertson

Roop

1999

Molecular Microbiology

174

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SummaryBrucella abortus is a facultative intracellular pathogen that causes abortion and infertility in domestic animals and a severe debilitating febrile illness in humans. The mechanisms that this highly successful intracellular pathogen uses to adapt to, and survive within, the harsh intracellular environment of the host macrophage are presently unknown. Maintenance of the stationary phase growth state has been proposed to be critical for the virulence of several mammalian pathogens, but analysis of this relationship for the brucellae has not been undertaken. In order to evaluate this relationship, we examined the in vitro and in vivo characteristics of an isogenic hfq mutant constructed from virulent Brucella abortus 2308. In Escherichia coli, the hfq gene product is an RNA-binding protein that participates in the regulation of stationary phase stress resistance, at least partly by enhancing translation of the stationary phase-speci®c sigma factor RpoS. As expected, the Brucella abortus hfq mutant, designated Hfq3, showed increased sensitivity to H 2 O 2, and decreased survival under acidic conditions (pH 4.0), during stationary phase growth compared with 2308. Hfq3 was also less able to withstand prolonged starvation than 2308. The Brucella abortus hfq mutant, unlike its parental strain 2308, fails to replicate in cultured murine macrophages, and is rapidly cleared from the spleens and livers of experimentally infected BALB/c mice. These ®ndings suggest that the Brucella abortus hfq gene product makes an essential contribution to pathogenesis in mice, probably by allowing the brucellae to adapt appropriately to the harsh environmental conditions encountered within the host macrophage.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The Brucella abortus host factor I (HF‐I) protein contributes to stress resistance during stationary phase and is a major determinant of virulence in mice

Robertson

Roop

1999

Molecular Microbiology

174

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“…Catalase deletion mutants of strains 2308 and strain 19 were constructed in a similar manner by gene replacement. Plasmid pCat5 (62), which contains the entire Brucella catalase coding sequence and flanking sequences, was doubly digested with BglII and PstI to remove 759 bp of the catalase coding region. The staggered ends were polished to blunt ends by incubation with the Klenow fragment of E. coli DNA polymerase I, and the neomycin-kanamycin resistance gene from Tn5 was inserted into the plasmid, effectively replacing the 3Ј half of the catalase gene in the plasmid (29).…”

Section: Resultsmentioning

confidence: 99%

“…Antibodies used for the Western blots were polyclonal rabbit antisera. Antisera for this study were developed in our laboratory and described in detail previously (10,16,27,29,62). The Western blot procedure used was based on that of Towbin et al (72).…”

Section: Methodsmentioning

confidence: 99%

Regulation of Brucella abortus Catalase

2000

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All aerobic organisms have mechanisms that protect against oxidative compounds. Catalase, peroxidase, superoxide dismutase, glutathione, and thioredoxin are widely distributed in many taxa and constitute elements of a nearly ubiquitous antioxidant metabolic strategy. Interestingly, the regulatory mechanisms that control these elements are rather different depending on the nature of the oxidative stress and the organism. Catalase is well documented to play an important role in protecting cells from oxidative stress. In particular, pathogenic bacteria seem to use this enzyme as a defensive tool against attack by the host. To investigate the significance of catalase in hostile environments, we made catalase deletion mutations in two different B. abortus strains and used two-dimensional gel analysis, survival tests, and adaptation experiments to explore the behavior and role of catalase under several oxidative stress conditions. These studies show that B. abortus strains that do not express catalase activity exhibit increased sensitivity to hydrogen peroxide. We also demonstrate that catalase expression is regulated in this species, and that preexposure to a sublethal concentration of hydrogen peroxide allows B. abortus to adapt so as to survive subsequent exposure to higher concentrations of hydrogen peroxide.

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“…Antibodies for the Western blots were polyclonal rabbit antisera. Antisera for this study were developed in our laboratory and described in detail previously (11,26,59). The procedure is based on Towbin et al (66).…”

Section: Western Blotmentioning

confidence: 99%

Regulation of Brucella abortus catalase as a defensive mechanism against oxidative stress

Kim

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Stinictiiral components of Brucella Pathogenesis of Brucella infection Oxidative stress in phagocytes Bacterial defensive mechanisms Catalase and superoxide dismutase Transcription factors of the LysR family 10 Rationale for this study References CHAPTER n. BRUCELLA ABORTUS PERIPLASMIC CATALASE IS REGULATED BY EXTERNAL HYDROGEN PEROXIDE 21 ABSTRACT INTRODUCTION 22 MATERIALS AND METHODS 24 Bacterial strains and medium 24 Construction of the deletion plasmid 24 Transformation of Brucella abortus 25 Halo assay (Hydrogen peroxide sensitivity assay) 25 2-D protein gel analysis 25 Western blot 26 Catalase assay 27 V Dot blot 27 RESUUrS Sensitivity of the catalase deletion mutant to HzCh 2-D gel analysis and identification of protein spots Response of Brucella abortus to oxidative stress 2-D gel analysis on Cu-Zn SOD deletion mutant 2-D gel analysis on catalase deletion mutant Enzyme activity Dot blot of catalase RNA DISCUSSION 31 REFERENCES CHAPTER in. OXYR REGULATES PERIPLASMIC CATALASE IN BRUCELLA ABORTUS

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Purification, characterization and cloning of a periplasmic catalase from B abortus and the role it plays in the pathogenesis of Brucella

Cited by 9 publications

References 83 publications

The Brucella abortus host factor I (HF‐I) protein contributes to stress resistance during stationary phase and is a major determinant of virulence in mice

The Brucella abortus host factor I (HF‐I) protein contributes to stress resistance during stationary phase and is a major determinant of virulence in mice

Regulation of Brucella abortus Catalase

Regulation of Brucella abortus catalase as a defensive mechanism against oxidative stress

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