Structural characterization of the active form of PerR: insights into the metal‐induced activation of PerR and Fur proteins for DNA binding

Jacquamet, Lilian; Traore, Daouda A K; Ferrer, Jean‐Luc; Proux, Olivier; Testemale, Denis; Hazemann, Jean‐Louis; Nazarenko, Elena; Ghazouani, Abdelnasser El; Caux‐Thang, Christelle; Duarte, Victor; Latour, Jean Marc

doi:10.1111/j.1365-2958.2009.06753.x

Cited by 103 publications

(162 citation statements)

References 46 publications

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“…3A). Metal binding to the sensor site promotes a change in PerR conformation from an extended to a more compact form that binds promoter DNA (5,6). The metal-binding-promoted change of PerR to the compact conformation is explained by the fact that H37 and H91, both of which participate directly as metal binding, are located in the two different domains of PerR; thus, these residues can help bring the two PerR domains in each monomer in close proximity and stabilize the compact form.…”

Section: Resultsmentioning

confidence: 99%

Oxidization without substrate unfolding triggers proteolysis of the peroxide-sensor, PerR

Ahn

Baker

2015

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

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Peroxide operon regulator (PerR) is a broadly conserved hydrogen peroxide sensor in bacteria, and oxidation of PerR at its regulatory metal-binding site is considered irreversible. Here, we tested whether this oxidation specifically targets PerR for proteolysis. We find that oxidizing conditions stimulate PerR degradation in vivo, and LonA is the principal AAA+ (ATPases associated with diverse cellular activities) protease that degrades PerR. Degradation of PerR by LonA is recapitulated in vitro, and biochemical dissection of this degradation reveals that the presence of regulatory metal and PerR-binding DNA dramatically extends the half-life of the protein. We identified a LonA-recognition site critical for oxidation-controlled PerR turnover. Key residues for LonA-interaction are exposed to solvent in PerR lacking metal, but are buried in the metal-bound form. Furthermore, one residue critical for Lon recognition is also essential for specific DNAbinding by PerR, thus explaining how both the metal and DNA ligands prevent PerR degradation. This ligand-controlled allosteric mechanism for protease recognition provides a compelling explanation for how the oxidation-induced conformational change in PerR triggers degradation. Interestingly, the critical residues recognized by LonA and exposed by oxidation do not function as a degron, because they are not sufficient to convert a nonsubstrate protein into a LonA substrate. Rather, these residues are a conformation-discriminator sequence, which must work together with other residues in PerR to evoke efficient degradation. This mechanism provides a useful example of how other proteins with only mild or localized oxidative damage can be targeted for degradation without the need for extensive oxidation-dependent protein denaturation.oxidative damage | proteolysis | degradation signals | Lon

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Section: Resultsmentioning

confidence: 99%

Oxidization without substrate unfolding triggers proteolysis of the peroxide-sensor, PerR

Ahn

Baker

2015

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

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“…Each PerR monomer contains a binding site for a structural Zn 2ϩ as well as a regulatory site that, in B. subtilis, binds either Fe 2ϩ or Mn 2ϩ (46,52) (Fig. 2A).…”

Section: Perrmentioning

confidence: 99%

Peroxide‐Sensing Transcriptional Regulators in Bacteria

Dubbs

Mongkolsuk

2016

Stress and Environmental Regulation of Gene Expression and Adaptation in Bacteria

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The ability to maintain intracellular concentrations of toxic reactive oxygen species (ROS) within safe limits is essential for all aerobic life forms. In bacteria, as well as other organisms, ROS are produced during the normal course of aerobic metabolism, necessitating the constitutive expression of ROS scavenging systems. However, bacteria can also experience transient high-level exposure to ROS derived either from external sources, such as the host defense response, or as a secondary effect of other seemingly unrelated environmental stresses. Consequently, transcriptional regulators have evolved to sense the levels of ROS and coordinate the appropriate oxidative stress response. Three well-studied examples of these are the peroxide responsive regulators OxyR, PerR, and OhrR. OxyR and PerR are sensors of primarily H 2 O 2 , while OhrR senses organic peroxide (ROOH) and sodium hypochlorite (NaOCl). OxyR and OhrR sense oxidants by means of the reversible oxidation of specific cysteine residues. In contrast, PerR senses H 2 O 2 via the Fe-catalyzed oxidation of histidine residues. These transcription regulators also influence complex biological phenomena, such as biofilm formation, the evasion of host immune responses, and antibiotic resistance via the direct regulation of specific proteins.

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“…The other resides in the Ddomain, constructed by either four conserved cysteines near the C terminus (C-site) or residues at the surface of the D-domain pointing toward the hinge region (D-site). Structural comparison between inactive apo-and metallated active forms of BsPerR suggested that regulatory metal binding causes DB-domains to swing around the hinge region with respect to the D-domain, from an open (swung-out) to closed (converged) conformations (22)(23)(24). This process leads to a model that the rearrangement of DBdomains by binding a regulatory metal in the hinge region is responsible for the allosteric transition of all Fur family regulators, from the inactive open conformation to the active closed one (23).…”

mentioning

confidence: 99%

“…The mechanisms of metal-specificity and metal-mediated activity modulation have been extensively investigated, revealing some common and specific principles among members of this family. Crystal structures of several metal-bound Fur regulators have been reported for iron-responsive Fur from Pseudomonas aeruginosa (PaFur) (18) and Vibrio cholerae (VcFur) (19), zincresponsive Zur from Mycobacterium tuberculosis (MtZur) (20), nickel-responsive Nur from S. coelicolor (ScNur) (21), and peroxide-sensing PerR from Bacillus subtilis (BsPerR) (22). All of the available structures demonstrate that the Fur family members are homodimeric and each monomer consists of an N-terminal DNAbinding (DB) domain, a C-terminal dimerization (D) domain, and a hinge loop between the two.…”

mentioning

confidence: 99%

Graded expression of zinc-responsive genes through two regulatory zinc-binding sites in Zur

Shin

Jung

et al. 2011

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

110

189

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Zinc is one of the essential transition metals in cells. Excess or lack of zinc is detrimental, and cells exploit highly sensitive zinc-binding regulators to achieve homeostasis. In this article, we present a crystal structure of active Zur from Streptomyces coelicolor with three zinc-binding sites (C-, M-, and D-sites). Mutations of the three sites differentially affected sporulation and transcription of target genes, such that C- and M-site mutations inhibited sporulation and derepressed all target genes examined, whereas D-site mutations did not affect sporulation and derepressed only a sensitive gene. Biochemical and spectroscopic analyses of representative metal site mutants revealed that the C-site serves a structural role, whereas the M- and D-sites regulate DNA-binding activity as an on-off switch and a fine-tuner, respectively. Consistent with differential effect of mutations on target genes, zinc chelation by TPEN derepressed some genes ( znuA, rpmF2 ) more sensitively than others ( rpmG2 , SCO7682) in vivo. Similar pattern of TPEN-sensitivity was observed for Zur-DNA complexes formed on different promoters in vitro. The sensitive promoters bound Zur with lower affinity than the less sensitive ones. EDTA-treated apo-Zur gained its DNA binding activity at different concentrations of added zinc for the two promoter groups, corresponding to free zinc concentrations of 4.5 × 10 −16 M and 7.9 × 10 −16 M for the less sensitive and sensitive promoters, respectively. The graded expression of target genes is a clever outcome of subtly modulating Zur-DNA binding affinities in response to zinc availability. It enables bacteria to detect metal depletion with improved sensitivity and optimize gene-expression pattern.

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Structural characterization of the active form of PerR: insights into the metal‐induced activation of PerR and Fur proteins for DNA binding

Cited by 103 publications

References 46 publications

Oxidization without substrate unfolding triggers proteolysis of the peroxide-sensor, PerR

Oxidization without substrate unfolding triggers proteolysis of the peroxide-sensor, PerR

Peroxide‐Sensing Transcriptional Regulators in Bacteria

Graded expression of zinc-responsive genes through two regulatory zinc-binding sites in Zur

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