6 research outputs found

    Mn(II) and Fe(II) modulate the binding activity of DR2539 <i>in vivo</i>.

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    <p>(A) Effects of divalent metals (50 ┬ÁM) on expression of pRAZH in <i>D. radiodurans</i>. Data shown are the means ┬▒ standard deviations of three independent experiments. (B) Effects of Mn(II) (squares) and Fe(II) (circles) on the expression of pRAZH in wild-type samples expressing DR2539. (C) Effects of Mn(II) (squares) and Fe(II) (circles) on the expression of pRAZH in the <i>dr2539</i> null mutant. (D) Rea-time PCR analysis of the <i>dr1709</i> gene expression using <i>dr0089</i> as internal control gene. Longitudinal axes indicate the change fold of <i>dr1709</i> mRNA relative to controls. Control cells were cultured in medium without Mn(II). *, <i>P</i><0.05 relative to control. The data are the means ┬▒ standard deviations of three independent experiments.</p

    DR2539 binds to the MntH promoter DNA fragment in a Mn(II)- and Fe(II)-dependent manner.

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    <p>(A) Schematic of <i>dr1709</i> promoter (p1709a and p1709b) DNA sequence region. The inverted repeat region is shown by the inverted arrows. (B) DR2539 binding to p1709b with increasing quantities of DR2539 and 25 ┬ÁM Mn(II). (C) and (D) EMSA analysis was performed using DR2539 and p1709b with increasing concentration of Mn(II) or Fe(II). RBS, ribosome binding site; Start, transcription start codon. p1709a and p1709b sequence regions are underlined by straight lines and dashed lines, respectively.</p

    DR0865 binds to the promoter of MntABC in an ion-dependent manner.

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    <p>(A) and (B) DR0865 binding to p2523 and p2284 as the concentration of DR0865 increased. (C) Wild-type R1, <i>dr2539</i> null mutant (<b>Δ</b><i>dr2539</i>), and <i>dr0865</i> null mutant (<b>Δ</b><i>dr0865</i>) were cultured on TGY plates overlaid with filter discs saturated with 1 M solution MnCl<sub>2</sub>. (D) The zone of inhibition was measured from edge of disc after three days. *, <i>P</i><0.05. Data represent the means±deviations of three independent experiments.</p

    Sequence alignment of the metal binding sites of DR2539 with other DxtR/MntR family members.

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    <p>ScaR (<i>Streptococcus gordonii</i>), SloR (<i>Streptococcus suis</i>), LCAS (<i>Lactobacillus casei</i>), SirR (<i>Corynebacterium glutamicum</i>), TroR (<i>Treponema pallidum</i>), TroR (<i>Treponema denticola</i>), IdeR (<i>Mycobacterium tuberculosis</i>), DtxR (<i>Corynebacterium diptheriae</i>), DR2539 (<i>Deinococcus radiodurans</i>), MntR (<i>Staphylococcus aureus</i>), MntR (<i>Escherichia coli</i>), and MntR (<i>Bacillus subtilis</i>). The sequences were aligned using the CLUSTAL W software. Residues shaded with black represent metal-binding sites that have been studied while residues shaded with grey represent predicted metal binding sites.</p

    His98 plays an important role in DNA binding activity of DR2539.

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    <p>(A) 10 ┬Ál cell dilution was dripped on the TGY plate to which 6 mM of Mn(II) had been added. The cells were cultured for 3 days. (B) H98Y mutant and wild-type DR2539 proteins were incubated with p1709b at different concentrations of Mn(II). (C) Quantification of the fluorescence intensity of binding bands was performed using ImageJ. *, <i>P</i><0.05.</p

    Proteomic insights into the functional basis for the response regulator DrRRA of <i>Deinococcus radiodurans</i>

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    <p><b>Purpose</b> To investigate the function basis of the recently discovered response regulator, <i>drRRA</i> (DNA damage response regulator A) in <i>Deinococcus radiodurans</i>, we compared the proteomic profile of the radiation-sensitive <i>drRRA</i> mutant with that of wild-type strain under both non-stress and gamma radiation treatment.</p> <p><b>Materials and methods</b> Total proteins of <i>D. radiodurans</i> cells were subjected to two-dimension electrophoresis. Protein spots in 2-Dimension gels were silver stained and scanned. Spots that changed significantly in expression levels were selected for mass spectrometry analysis. Seven genes encoding representative proteins were knocked out for stress resistance analysis.</p> <p><b>Results</b> A total of 52 proteins displayed significant expression level changes at least 1.5-fold in the mutant relative to wild-type strain under non-stress conditions, with 31 repressed and 21 induced proteins, which might affect the cell response of <i>D. radiodurans</i> to gamma radiation. The proteins were distributed into functional groups including stress response, metabolism, and function unknown. Disruptions of several altered proteins including DRA0259 (Catalase E) and DR1538 (Osmotically inducible protein C), reduced the antioxidant activity of <i>D. radiodurans</i>.</p> <p><b>Conclusion</b> Combined with our previous result of transcriptional profile, we further confirmed that inactivation of DrRRA affects the expression of various stress response systems.</p