Crystal structure of the cyanobacterial metallothionein repressor SmtB: a model for metalloregulatory proteins 1 1Edited by D. Rees

Cook, William J.; Kar, S.; Taylor, Kenneth B.; Hall, Leo M.

doi:10.1006/jmbi.1997.1443

Cited by 130 publications

(169 citation statements)

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“…The N terminus contains an HTH DNA-binding motif common among most of the metalloregulatory proteins (Lam et al, 1994 ;Cook et al, 1998 ;Pohl et al, 1999). These secondary-structure-associated features suggest that S. aureus Fur is a metal-responsive regulatory protein.…”

Section: Resultsmentioning

confidence: 99%

Molecular characterization of the ferric-uptake regulator, Fur, from Staphylococcus aureus The GenBank accession numbers for the S. aureus fur gene and fhu operon reported in this paper are AF118839 and AF132117, respectively.

et al. 2000

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Iron is an essential nutrient for the survival and pathogenesis of bacteria, but relatively little is known regarding its transport and regulation in staphylococci. Based on the known sequences of ferric-uptake regulatory (fur) genes from several Gram-positive and Gram-negative bacteria, a fragment containing the fur homologue was cloned from a genomic library of Staphylococcus aureus RN450. Nucleotide sequence analysis of this fragment revealed the presence of a 447 bp ORF that encodes a putative 149 aa polypeptide with an apparent molecular mass of 17 kDa. A putative ferrichrome-uptake (fhu) operon, containing the conserved Fur-binding sequences (Fur box) in the promoter region, was also cloned from the same S. aureus library. To characterize the impact of Fur on the fhu operon, fur was cloned, overexpressed as a His-tagged protein and purified by Ni 2M -affinity column chromatography. The recombinant protein was digested with enterokinase to remove the His tag. Electrophoretic mobility-shift assays indicated that Fur binds to the promoter region of the fhu operon in the presence of divalent cations. Fur also interacted with the promoter region of the recently reported sir operon that has been proposed to constitute a siderophore-transport system in S. aureus. The DNase I-protection assay revealed that Fur specifically binds to the Fur box located in the promoter region of the fhu operon. The primer-extension reaction indicated that the transcription-start site of the fhu operon was located inside the Fur box. S. aureus fur partially complemented a fur N mutation in Bacillus subtilis. The data suggest that Fur regulates iron-transport processes in S. aureus.

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Section: Resultsmentioning

confidence: 99%

Molecular characterization of the ferric-uptake regulator, Fur, from Staphylococcus aureus The GenBank accession numbers for the S. aureus fur gene and fhu operon reported in this paper are AF118839 and AF132117, respectively.

et al. 2000

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“…The crystal structure of CzrA and SmtB in the apo and the Zn(II)-bound states have been solved (8). Although the structures of CzrA were found to be very similar in these two states, SmtB revealed measurable differences in quaternary structure in which the apo form adopted a comparatively ''flat'' conformation not well suited to interact with canonical B-form DNA (8,15). The structural and thermodynamic underpinnings of metalloregulation for any member of the ubiquitous ArsR family remains poorly understood due to a lack of detailed insight for the DNA operator-bound state (3).…”

Section: Staphylococcus Aureusmentioning

confidence: 99%

“…Fig. 3A shows { 1 H}- 15 N hNOE values for the DNA-bound CzrA. Amide bonds (NH) that are immobile over this time scale show NOE values greater than or equal to 0.8, whereas those NH bonds that experience internal motions in subnanosecond time scale show lower hNOE values.…”

Section: Induced Flexibility In the Allosteric Metal-binding Sites Inmentioning

confidence: 99%

“…Backbone and stereospecific methyl side chain resonance assignments were carried out as reported elsewhere (36). Backbone torsional angles ( / ) were derived from backbone 1 H, 15 N, and 13 C chemical shifts using the program TALOS (38). Distance constraints for both intramolecular and intersubunit contacts were derived from 3D 13 C/ 13 C-separated methyl NOE and TROSY-version of the 3D 15 N-separated NOE experiment.…”

Section: Nmr Experiments and Structurementioning

confidence: 99%

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Solution structure of a paradigm ArsR family zinc sensor in the DNA-bound state

Arunkumar

Campanello

Giedroc

2009

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

180

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Staphylococcus aureusCzrA is a zinc-dependent transcriptional repressor from the ubiquitous ArsR family of metal sensor proteins. Zn(II) binds to a pair of intersubunit C-terminal ␣5-sensing sites, some 15 Å distant from the DNA-binding interface, and allosterically inhibits DNA binding. This regulation is characterized by a large allosteric coupling free energy (⌬Gc) of approximately ؉6 kcal mol ؊1 , the molecular origin of which is poorly understood. Here, we report the solution quaternary structure of homodimeric CzrA bound to a palindromic 28-bp czr operator, a structure that provides an opportunity to compare the two allosteric ''end'' states of an ArsR family sensor. Zn ( M etal ion homeostasis is a complex process that involves maintaining a delicate balance between metal uptake/ efflux and other metal storage systems to meet the needs of the cell (1, 2). This process is tightly regulated at the level of transcription by means of specific metal-dependent transcriptional regulators that respond to changes in metal ion concentrations in the host environment (3). In Staphylococcus aureus, the czr operon encodes a CDF antiporter, CzrB, a homolog of Escherichia coli zinc transporter YiiP that confers resistance to Zn(II) and Co(II) (4, 5), and the metal-regulated repressor, CzrA (6, 7). CzrA binds Zn(II) with picomolar affinity and strong negative homotropic cooperativity (8, 9) and is thought to undergo a conformational change that alleviates transcriptional repression of the resistance gene czrB.CzrA belongs to the ubiquitous ArsR (or ArsR/SmtB) family of metalloregulators found in many bacterial genomes that sense a wide variety of metals including biologically essential metals as well as toxic metal pollutants (10, 11). Members of this family appear to adopt a common winged helix-turn-helix homodimeric fold, but have evolved physically and structurally distinct pairs of allosteric metal-sensing sites (2, 12). These sites are thought to have arisen as a result of convergent evolution due to evolutionary pressures (12), a finding consistent with the ''rule of varied allosteric control'' in which protein families evolve seemingly random allosteric control pathways (13). CzrA and its homolog SmtB in Synechococcus (14) are ␣5 sensors that bind Zn(II) ions in two rotationally symmetric tetrahedral coordination sites formed by pairs of metal ligands derived from the ␣5 helix of each subunit (8). The crystal structure of CzrA and SmtB in the apo and the Zn(II)-bound states have been solved (8). Although the structures of CzrA were found to be very similar in these two states, SmtB revealed measurable differences in quaternary structure in which the apo form adopted a comparatively ''flat'' conformation not well suited to interact with canonical B-form DNA (8,15). The structural and thermodynamic underpinnings of metalloregulation for any member of the ubiquitous ArsR family remains poorly understood due to a lack of detailed insight for the DNA operator-bound state (3).We report here the NMR solution structure of ...

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“…All of these proteins form a biological dimer, and the structure of PA1607 is consistent with this, though the formation of the dimer is altered in PA1607. Additionally, some proteins with similar structure have been proposed to have a metal-binding site (Cook et al 1998). This structure suggests that the metal-binding site is not conserved in PA1607.…”

mentioning

confidence: 99%

The X‐ray crystal structure of PA1607 from Pseudomonas aureginosa at 1.9 Å resolution—a putative transcription factor

Sieminska

Savchenko

et al. 2007

Protein Science

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The structure of the PA1607 protein from Pseudomonas aureginosa was determined at 1.85 Å resolution using the Se-Met multiwavelength anomalous diffraction (MAD) technique. PA1607 forms a dimer and adopts a winged-helix motif similar to the MarR family of transcription regulators, though it has an unusual dimerization profile. The DNA-binding regions and a putative metal-binding site are not conserved in PA1607.

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Crystal structure of the cyanobacterial metallothionein repressor SmtB: a model for metalloregulatory proteins 1 1Edited by D. Rees

Cited by 130 publications

References 40 publications

Molecular characterization of the ferric-uptake regulator, Fur, from Staphylococcus aureus The GenBank accession numbers for the S. aureus fur gene and fhu operon reported in this paper are AF118839 and AF132117, respectively.

Molecular characterization of the ferric-uptake regulator, Fur, from Staphylococcus aureus The GenBank accession numbers for the S. aureus fur gene and fhu operon reported in this paper are AF118839 and AF132117, respectively.

Solution structure of a paradigm ArsR family zinc sensor in the DNA-bound state

The X‐ray crystal structure of PA1607 from Pseudomonas aureginosa at 1.9 Å resolution—a putative transcription factor

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