JOURNAL OF BACTERIOLOGY, Feb. 2003, p. 1010–1017 0021-9193/03/$08.00 0 DOI: 10.1128/JB.185.3.1010–1017.2003 Copyright © 2003, American Society for Microbiology. All Rights Reserved.
Vol. 185, No. 3
The Iron-Binding Protein Dps Confers Hydrogen Peroxide Stress Resistance to Campylobacter jejuni
Takahiko Ishikawa,1,2 Yoshimitsu Mizunoe,1* Shun-ichiro Kawabata,3 Akemi Takade,1 Mine Harada,2Sun Nyunt Wai,1,4 and Shin-ichi Yoshida1
Department of Bacteriology1 and Department of Medicine and Biosystemic Science, Internal Medicine,2 Faculty of Medical Sciences, and Department of Biology, Faculty of Sciences,3 Kyushu University, Fukuoka 812-8582, Japan, and Department of Molecular Biology, Umea University, S901 87 Umea, Sweden4 ˚ ˚
Received 26 September 2002/Accepted 15 November 2002We identiﬁed and characterized the iron-binding protein Dps from Campylobacter jejuni. Electron microscopic analysis of this protein revealed a spherical structure of 8.5 nm in diameter, with an electron-dense core similar to those of other proteins of the Dps (DNA-binding protein from starved cells) family. Cloning and sequencing of the Dps-encoding gene (dps) revealed that a 450-bp open readingframe (ORF) encoded a protein of 150 amino acids with a calculated molecular mass of 17,332 Da. Amino acid sequence comparison indicated a high similarity between C. jejuni Dps and other Dps family proteins. In C. jejuni Dps, there are iron-binding motifs, as reported in other Dps family proteins. C. jejuni Dps bound up to 40 atoms of iron per monomer, whereas it did not appear to bind DNA. Anisogenic dps-deﬁcient mutant was more vulnerable to hydrogen peroxide than its parental strain, as judged by growth inhibition tests. The iron chelator Desferal restored the resistance of the Dps-deﬁcient mutant to hydrogen peroxide, suggesting that this iron-binding protein prevented generation of hydroxyl radicals via the Fenton reaction. Dps was constitutively expressed during both exponential andstationary phase, and no induction was observed when the cells were exposed to H2O2 or grown under iron-supplemented or iron-restricted conditions. On the basis of these data, we propose that this iron-binding protein in C. jejuni plays an important role in protection against hydrogen peroxide stress by sequestering intracellular free iron and is expressed constitutively to cope with the harmfuleffect of hydrogen peroxide stress on this microaerophilic organism without delay. The microaerophilic gram-negative bacterium Campylobacter jejuni is a major cause of bacterial diarrhea in both developed and developing countries. Infection with C. jejuni usually causes an acute, self-limited gastroenteritis characterized by diarrhea, fever, nausea, and abdominal cramps. The clinical aspects rangefrom watery to severely bloody diarrhea (3, 26). In addition to acute gastrointestinal disease, infection with C. jejuni is recognized as the most important trigger of GuillainBarre syndrome (2). Despite its great importance in human ´ health and economy, little is known of its virulence determinants and mechanisms of survival in the environment. In their natural environment, whether in thegastrointestinal tracts of mammalian hosts or in the external environment during transmission, it is likely that C. jejuni cells are faced with growth-limiting or potentially lethal conditions, such as iron limitation and oxidative stresses. Iron is an essential nutrient for most organisms. It is required as a cofactor by several enzymes and as a catalyst in electron transport processes. However, inthe presence of oxygen, iron is able to generate oxygen radicals that can damage DNA, proteins, and membrane lipids (19, 23, 32). Iron homeostasis therefore has to be carefully balanced, and this is achieved by tightly controlling its uptake, metabolism, and storage (12). The intracellular amount of free ferrous iron is controlled mainly by the iron-binding protein ferritin, which
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