HucR, a Novel Uric Acid-responsive Member of the MarR Family of Transcriptional Regulators from Deinococcus radiodurans

Wilkinson, Steven; Grove, Anne

doi:10.1074/jbc.m405586200

Cited by 102 publications

(132 citation statements)

References 57 publications

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“…[39][40][41] Complex formation with DNA is required for MexR to form a jointly folded entity with a single, highly cooperative, unfolding transition. In this context, it is interesting that the structural homologue HucR, which is prefolded for DNAbinding, displays a single cooperative melting transition at 51 C already in the absence of DNA; 14,42 this T m is close to what is observed for the dimer region in MexR both in its apo and DNA-bound states.…”

Section: Discussionmentioning

confidence: 71%

Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance

et al. 2010

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The self-assembling MexA-MexB-OprM efflux pump system, encoded by the mexO operon, contributes to facile resistance of Pseudomonas aeruginosa by actively extruding multiple antimicrobials. MexR negatively regulates the mexO operon, comprising two adjacent MexR binding sites, and is as such highly targeted by mutations that confer multidrug resistance (MDR). To understand how MDR mutations impair MexR function, we studied MexR-wt as well as a selected set of MDR single mutants distant from the proposed DNA-binding helix. Although DNA affinity and MexA-MexB-OprM repression were both drastically impaired in the selected MexR-MDR mutants, MexR-wt bound its two binding sites in the mexO with high affinity as a dimer. In the MexR-MDR mutants, secondary structure content and oligomerization properties were very similar to MexR-wt despite their lack of DNA binding. Despite this, the MexR-MDR mutants showed highly varying stabilities compared with MexR-wt, suggesting disturbed critical interdomain contacts, because mutations in the DNA-binding domains affected the stability of the dimer region and vice versa. Furthermore, significant ANS binding to MexR-wt in both free and DNA-bound states, together with increased ANS binding in all studied mutants, suggest that a hydrophobic cavity in the dimer region already shown to be involved in regulatory binding is enlarged by MDR mutations. Taken together, we propose that the biophysical MexR properties that are targeted by MDR mutations-stability, domain interactions, and internal hydrophobic surfaces-are also critical for the regulation of MexR DNA binding.

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Section: Discussionmentioning

confidence: 71%

Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance

et al. 2010

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“…The data were calculated from three independent experiments and are shown as the mean+SD from the nine samples (Lin et al, 2006). The pH of the modified LB broth, which had been supplemented with different concentrations of urate (dissolved in 1 M NaOH), was adjusted to 7.5 with HCl as described in Wilkinson & Grove (2004). NaOH and HCl were also added to modify the no-urate control LB.…”

Section: Methodsmentioning

confidence: 99%

PecS regulates the urate-responsive expression of type 1 fimbriae in Klebsiella pneumoniae CG43

et al. 2015

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In the Klebsiella pneumoniae CG43 genome, the divergently transcribed genes coding for PecS, the MarR-type transcription factor, and PecM, the drug metabolite transporter, are located between the type 1 and type 3 fimbrial gene clusters. The intergenic sequence pecO between pecS and pecM contains three putative PecS binding sites and a CpxR box. Electrophoretic mobility shift assay revealed that the recombinant PecS and CpxR could specifically bind to the pecO sequence, and the specific interaction of PecS and pecO could be attenuated by urate. The expression of pecS and pecM was negatively regulated by CpxAR and PecS, and was inducible by exogenous urate in the absence of cpxAR. Compared with CG43S3DcpxAR, the derived mutants CG43S3DcpxARDpecS and CG43S3DcpxARDpecSDpecM exerted similar levels of sensitivity to H 2 O 2 or paraquat, but higher levels of mannose-sensitive yeast agglutination activity and FimA production. The promoter activity and transcript levels of fimA in CG43S3DcpxAR were also increased by deleting pecS. However, no binding activity between PecS and the fimA promoter could be observed. Nevertheless, PecS deletion could reduce the expression of the global regulator HNS and release the negative effect of HNS on FimA expression. In CG43S3DcpxAR, the expression of FimA as well as PecS was inducible by urate, whilst urate-induced FimA expression was inhibited by the deletion of pecS. Taken together, we propose that K. pneumoniae PecS indirectly and negatively regulates the expression of type 1 fimbriae, and the regulation is urateinducible in the absence of CpxAR.

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“…The engineered cells were encapsulated in mice for the treatment of gout and tumor lysis syndrome, which results from the formation of urate crystals in the joints and kidneys, respectively. This system was able to restore urate homeostasis through detection of high urate levels and concomitant derepression of a fungal urate oxidase, a urate-metabolizing enzyme that has been lost in humans (65,66). Efforts continue to be directed toward engineering dynamic mammalian circuits, including schemes that combine drug-based and gene-based approaches.…”

Section: In Vivo Diagnosticsmentioning

confidence: 99%

Synthetic biology devices for in vitro and in vivo diagnostics

Slomovic

Pardee

Collins

2015

Proc. Natl. Acad. Sci. U.S.A.

287

219

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There is a growing need to enhance our capabilities in medical and environmental diagnostics. Synthetic biologists have begun to focus their biomolecular engineering approaches toward this goal, offering promising results that could lead to the development of new classes of inexpensive, rapidly deployable diagnostics. Many conventional diagnostics rely on antibody-based platforms that, although exquisitely sensitive, are slow and costly to generate and cannot readily confront rapidly emerging pathogens or be applied to orphan diseases. Synthetic biology, with its rational and short design-to-production cycles, has the potential to overcome many of these limitations. Synthetic biology devices, such as engineered gene circuits, bring new capabilities to molecular diagnostics, expanding the molecular detection palette, creating dynamic sensors, and untethering reactions from laboratory equipment. The field is also beginning to move toward in vivo diagnostics, which could provide near real-time surveillance of multiple pathological conditions. Here, we describe current efforts in synthetic biology, focusing on the translation of promising technologies into pragmatic diagnostic tools and platforms.synthetic biology | diagnostics | biosensing | synthetic gene networks | nanobiotechnology

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HucR, a Novel Uric Acid-responsive Member of the MarR Family of Transcriptional Regulators from Deinococcus radiodurans

Cited by 102 publications

References 57 publications

Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance

Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance

PecS regulates the urate-responsive expression of type 1 fimbriae in Klebsiella pneumoniae CG43

Synthetic biology devices for in vitro and in vivo diagnostics

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