11 research outputs found

    Growth curves of <i>D</i>. <i>radiodurans</i> strains.

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    <p>(A) Wild-type and four CRP mutants were individually grown in TGY medium, and the OD<sub>600</sub> was measured every 2 h. (B) Growth rates of the wild-type, <i>dr0997</i> mutant, and the <i>dr0997</i> mutant complemented with the <i>dr0997</i> gene (<i>dr0997 mutant</i> Cwt) were recorded every 2 h.</p

    Effect of <i>dr0997</i> disruption on the anti-oxidant ability of <i>D</i>. <i>radiodurans</i>.

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    <p>(A) Total antioxidant capacity of the wild-type R1, <i>dr0997</i> mutant, and <i>dr0997</i> mutant complemented with the <i>dr0997</i> gene (<i>dr0997 mutant</i> Cwt) strains after exposure to 30 mM H<sub>2</sub>O<sub>2</sub> treatment. (B) Catalase activities after 30 mM H<sub>2</sub>O<sub>2</sub> treatment. Data represent the average and standard deviations of three independent experiments.</p

    Survival of CRP mutants exposed to MMC and UV radiation.

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    <p>(A) Survival of CRP mutants and the wild-type strain after exposure to 5 ╬╝g/ml MMC for different periods of time. (B) Survival of the <i>dr0997</i> mutant and the <i>dr0997</i> mutant complemented with the <i>dr0997</i> gene (<i>dr0997 mutant</i> Cwt) after MMC exposure. (C) Survival of the CRP mutants and the wild-type strain after exposure to UV radiation. (D) Survival curves of the wild-type, <i>dr0997</i> mutant, and <i>dr0997 mutant</i> Cwt strains after exposure to UV radiation (0 to 500 Jm<sup>Ôłĺ2</sup>). Data represent the averages and standard deviations of three independent experiments.</p

    Alignment of CRP homologs from different organisms.

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    <p>ClustalX software was used to align the amino acid sequence of <i>D</i>. <i>radiodurans</i> R1 DR0997, DR0834, DR1646, and DR2362, <i>M</i>. <i>tuberculosis</i> RV3676, <i>T</i>. <i>thermophilus</i> HB8 TTHA1357, and <i>E</i>. <i>coli</i> CRP. The conservative residues Gly71, Glu72, Arg82 and Thr172 (dotted frames) of <i>E</i>. <i>coli</i> CRP are required for cAMP binding, while Arg180 and Glu181 (solid frames) are required for DNA binding, based on the crystal structure of <i>E</i>. <i>coli</i> CRP.</p

    Cyclic AMP Receptor Protein Acts as a Transcription Regulator in Response to Stresses in <i>Deinococcus radiodurans</i>

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    <div><p>The cyclic AMP receptor protein family of transcription factors regulates various metabolic pathways in bacteria, and also play roles in response to environmental changes. Here, we identify four homologs of the CRP family in <i>Deinococcus radiodurans</i>, one of which tolerates extremely high levels of oxidative stress and DNA-damaging reagents. Transcriptional levels of CRP were increased under hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) treatment during the stationary growth phase, indicating that CRPs function in response to oxidative stress. By constructing all CRP single knockout mutants, we found that the <i>dr0997</i> mutant showed the lowest tolerance toward H<sub>2</sub>O<sub>2</sub>, ultraviolet radiation, ionizing radiation, and mitomycin C, while the phenotypes of the <i>dr2362</i>, <i>dr0834</i>, and <i>dr1646</i> mutants showed slight or no significant differences from those of the wild-type strain. Taking advantage of the conservation of the CRP-binding site in many bacteria, we found that transcription of 18 genes, including genes encoding chromosome-partitioning protein (<i>dr0998</i>), Lon proteases (<i>dr0349</i> and <i>dr1974</i>), NADH-quinone oxidoreductase (<i>dr1506</i>), thiosulfate sulfurtransferase (<i>dr2531</i>), the DNA repair protein UvsE <i>(dr1819</i>), PprA (<i>dra0346</i>), and RecN (<i>dr1447</i>), are directly regulated by DR0997. Quantitative real-time polymerase chain reaction (qRT-PCR) analyses showed that certain genes involved in anti-oxidative responses, DNA repair, and various cellular pathways are transcriptionally attenuated in the <i>dr0997</i> mutant. Interestingly, DR0997 also regulate the transcriptional levels of all CRP genes in this bacterium. These data suggest that DR0997 contributes to the extreme stress resistance of <i>D</i>. <i>radiodurans</i> via its regulatory role in multiple cellular pathways, such as anti-oxidation and DNA repair pathways.</p></div

    Survival curves of <i>D</i>. <i>radiodurans</i> strains exposed to 50 mM H<sub>2</sub>O<sub>2</sub>.

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    <p>(A) Wild-type and CRP mutants exposed to 50 mM H<sub>2</sub>O<sub>2</sub> for 20 min, and then plated on TGY agar plates. (B) Wild-type, <i>dr0997</i> mutant, and the <i>dr0997</i> mutant complemented with the <i>dr0997</i> gene (<i>dr0997 mutant</i> Cwt) were exposed to 50 mM H<sub>2</sub>O<sub>2</sub> for different periods of time. Data represent the averages and standard deviations of three independent experiments.</p
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