Elucidation of Primary (α3N) and Vestigial (α5) Heavy Metal-binding Sites in Staphylococcus aureus pI258 CadC: Evolutionary Implications for Metal Ion Selectivity of ArsR/SmtB Metal Sensor Proteins

Busenlehner, Laura S.; Weng, Tsu Chien; Penner‐Hahn, James E.; Giedroc, David P.

doi:10.1016/s0022-2836(02)00299-1

Cited by 108 publications

(256 citation statements)

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“…It is interesting that Fur and DtxR family members exhibit metal affinities generally much weaker than members of MerR and ArsR/SmtB families, the latter of which bind cognate ions in the 10 -9 to 10 -21 M range (8,37,40,(52)(53)(54)(55)(56)(57). One explanation for this apparent trend may come from considering the function of these metalloregulatory protein families.…”

Section: Metal Affinities Of Mntr Versus Other Metalloregulatory Protmentioning

confidence: 99%

“…Ni 2+ may also fail to form a dinuclear site, or the geometry of such a site with this metal ion may be significantly different, which again fails to produce the requisite allosteric change in MntR. Overall, we would concur with the hypothesis of Glasfeld et al (15) that the geometry and stoichiometry (dinuclear) of metal binding to MntR is more significant to the mechanism of activation than metal ion affinity.It is interesting that Fur and DtxR family members exhibit metal affinities generally much weaker than members of MerR and ArsR/SmtB families, the latter of which bind cognate ions in the 10 -9 to 10 -21 M range (8,37,40,(52)(53)(54)(55)(56)(57) (1 mM) and 40 times higher than that of Cd 2+ (0.5 mM) (63). Furthermore, in several Bacillus species, the minimal Mn 2+ concentration required for normal vegetative growth is ∼10 -7 M, 10 -6 -10 -3 M for production of secondary metabolites, and 10 -4 -10 -3 M for cultural longevity (64).…”

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Metal Binding Studies and EPR Spectroscopy of the Manganese Transport Regulator MntR

et al. 2006

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Manganese transport regulator (MntR) is a member of the diphtheria toxin repressor (DtxR) family of transcription factors that is responsible for manganese homeostasis in [4295][4296][4297][4298][4299][4300][4301][4302][4303], and generally follow the Irving-Williams series. Direct detection of the dinuclear Mn 2+ site in MntR with EPR spectroscopy is presented, and the exchange interaction was determined, J = -0.2 cm -1 . This value is lower in magnitude than most known dinuclear Mn 2+ sites in proteins and synthetic complexes and is consistent with a dinuclear Mn 2+ site with a longer Mn···Mn distance (4.4 Å) observed in some of the available crystal structures. MntR is found to have a surprisingly low binding affinity (∼160 μM) for its cognate metal ion Mn 2+ . Moreover, the results of DNA binding studies in the presence of limiting metal ion concentrations were found to be consistent with the measured metal-binding constants. The metalbinding affinities of MntR reported here help to elucidate the regulatory mechanism of this metaldependent transcription factor.Bacteria handle the delicate issue of metal ion homeostasis using a class of transcription factors known as metalloregulatory proteins (metalloregulators). In some systems, these metal-sensing proteins are involved in mediating the removal of toxic metals, while in other systems they are central to maintaining the required levels of essential metals. To date, a large number of metalloregulatory proteins have been identified that respond to a variety of metal ions (1-3). The subject of how metal binding is translated into an ability to control transcription via a † This work was supported by the University of California, San Diego, the Hellman Family fund, a Cottrell Scholar Award from the Research Corporation (S.M.C.), and the National Institutes of Health GM077387 (M.P.H.). * Author to whom correspondence should be adddressed. (M.P.H.) Telephone: (412) 268-1058; fax: (412) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript metalloregulator has become a prominent subject of investigation (4-10). Indeed, many metalloregulators are reported to bind several metal ions in vitro, while only eliciting a specific transcriptional response when they are bound to the cognate metal in vivo (5)(6)(7)9). This observation naturally leads to the question: how does a metalloregulator selectively respond to its cognate metal ion as opposed to other available metal ion activators? The ability of a metalloregulator to respond selectively to a metal ion may depend on several factors, including the availability of the requisite metal ion, the binding affinity for the metal ion, the charge on the metal ion, and the coordination geometry/number assumed by the metal ion upon binding. Determining which of these factors are most important for a given metalloregulator is essential for gaining a better understanding of how these proteins elicit transcriptional control.The manganese transport regulator MntR 1 is found in Bacillus subtilis and is ...

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Section: Metal Affinities Of Mntr Versus Other Metalloregulatory Protmentioning

confidence: 99%

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Metal Binding Studies and EPR Spectroscopy of the Manganese Transport Regulator MntR

et al. 2006

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“…As the concentration of metal ion increases, the effector-binding sites of the regulators become occupied eliciting a conformational change that weakens the affinity for the O͞P region, allowing transcription to proceed. Members of the SmtB͞ArsR family include: As(III), Sb(III), Bi(III)-responsive ArsR (7), Zn(II)-responsive SmtB (8), Cd(II), Pb(II), Bi(III)-responsive CadC (9)(10)(11), Zn(II)-responsive ZiaR (12), Co(II), Zn(II)-responsive CzrA (13,14), and, most recently, Ni(II), Co(II)-responsive NmtR (15).…”

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“…Comparative structural and spectroscopic studies of six SmtB͞ ArsR family members reveal that individual members are characterized by one or both of two structurally distinct metal coordination sites (6,11,(15)(16)(17)(18)(19)(20). These two metal sites are designated ␣3N (or ␣3) and ␣5 (or ␣5C), named for the location of the metal-binding ligands within the known or predicted secondary structure of individual family members.…”

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“…All fluorescence anisotropy experiments were carried out with an SLM 4800 (SLM-Aminco, Urbana, IL) spectrofluorometer operating in the steady-state mode fitted with Glan-Thompson (SLM Instruments, Urbana, IL) polarizers in the L format (11). The 57-mer double-stranded czr O͞P oligonucleotide (Operon Technologies, Alameda, CA) used was fluorescein-labeled on one 5Ј-end with the sequence given below.…”

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Structural elements of metal selectivity in metal sensor proteins

Pennella

Shokes

Cosper

et al. 2003

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

108

270

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Staphylococcus aureusA bout one-third of all proteins exploit specific metal ions to assist in macromolecular folding and͞or function at the active site of metalloenzymes (1). All cells restrict the number of bioavailable metal atoms to avoid any excess that would otherwise compete with native metal ion sites that do not support biological activity (2). Essentially all cell types contain intracellular metal sensors that detect surplus metal ions and control the expression of genes encoding proteins that expel or sequester the extra ions (3). For some metals and some cell types, a complementary set of sensors detect deficiency and regulate genes encoding proteins that acquire more of the required ions (4, 5). It is currently poorly understood how such metal-sensing metalloregulators accurately discriminate between various metal ions.SmtB͞ArsR-family regulators are ubiquitous in bacterial genomes and bind to the operator͞promoter (O͞P) regions of gene(s) encoding proteins involved in metal export or sequestration, repressing transcription (for a review, see ref. 6). As the concentration of metal ion increases, the effector-binding sites of the regulators become occupied eliciting a conformational change that weakens the affinity for the O͞P region, allowing transcription to proceed. Members of the SmtB͞ArsR family include: As(III), Sb(III), Bi(III)-responsive ArsR (7), Zn(II)-responsive SmtB (8), Cd(II), Pb(II), Bi(III)-responsive CadC (9-11), Zn(II)-responsive ZiaR (12), Co(II), Zn(II)-responsive CzrA (13,14), and, most recently, Ni(II), Co(II)-responsive NmtR (15).Comparative structural and spectroscopic studies of six SmtB͞ ArsR family members reveal that individual members are characterized by one or both of two structurally distinct metal coordination sites (6, 11, 15-20). These two metal sites are designated ␣3N (or ␣3) and ␣5 (or ␣5C), named for the location of the metal-binding ligands within the known or predicted secondary structure of individual family members. The coordination environment and precise ligand set of the ␣3, ␣3N, and͞or ␣5, ␣5C sites in the different SmtB͞ArsR proteins differ and are presumed to contribute toward metal selectivity. A sequence comparison for proteins discussed herein is shown in Fig. 1 and highlights these sites.Here we report insights gained from the study of two additional family members, Staphylococcus aureus CzrA and Mycobacterium tuberculosis NmtR. CzrA and NmtR share 30% sequence identity and a high degree of similarity (60%) yet respond to distinct but partially overlapping metal profiles in vivo. S. aureus CzrA is a Co(II)͞Zn(II)-specific sensor that regulates the expression of the czr operon, which encodes a Co(II)͞ Zn(II)-facilitated pump, CzrB, that effluxes metal out of the cell (13, 14). Electromobility-shift assays and in vivo expression studies indicate that Zn(II) is the strongest inducer of CzrA regulation, with Co(II) also capable of regulation but only at higher concentrations than Zn(II). Other metals, including Ni(II), have little to no effect on derep...

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Zinc:DNA‐Binding Proteins

Pennella¹,

Giedroc²

2005

Encyclopedia of Inorganic Chemistry

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Zinc‐containing nucleic acid‐binding proteins are ubiquitous in nature and interact with all types of nucleic acids, including ssDNA, duplex DNA, and RNA. The primary role played by these Zn(II) coordination complexes is to provide local or global stability to these proteins that function in transcriptional regulation, DNA metabolism, and nucleic acid packaging. These proteins are now broadly designated as zinc finger (zf) proteins. Six structural classes of zf proteins are discussed here that are distinguished from one another on the basis of the secondary and tertiary structure of the zf domain, the nature of the ligating side chains (Cys and/or His), and the linear spacing of amino acid ligands in the primary structure. These structural zinc sites are compared with metalloregulatory zinc coordination complexes found in metal‐sensing transcriptional regulatory proteins that control zinc homeostasis in bacteria (e.g. E. coli ZurR and ZntR; S. aureus CzrA; Synechococcus SmtB), lower eukaryotes ( S. cerevisiae Zap1), and mammalian cells (MTF‐1). Structural and functional comparisons of the bacterial zinc sensors with homologous sensors specific for other transition metal ions (e.g. Cu, Ni, Mn) and xenobiotics (e.g. Cd, Pb, and Hg) suggest that coordination number and therefore geometry are primary determinants of metal selectivity of metal sensor proteins in vivo.

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Elucidation of Primary (α3N) and Vestigial (α5) Heavy Metal-binding Sites in Staphylococcus aureus pI258 CadC: Evolutionary Implications for Metal Ion Selectivity of ArsR/SmtB Metal Sensor Proteins

Cited by 108 publications

References 38 publications

Metal Binding Studies and EPR Spectroscopy of the Manganese Transport Regulator MntR

Metal Binding Studies and EPR Spectroscopy of the Manganese Transport Regulator MntR

Structural elements of metal selectivity in metal sensor proteins

Zinc:DNA‐Binding Proteins

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