Control of Expression of a Periplasmic Nickel Efflux Pump by Periplasmic Nickel Concentrations

Grass, Gregor; Fricke, Beate; Nies, Dietrich H.

doi:10.1007/s10534-005-3718-6

Cited by 64 publications

(78 citation statements)

References 52 publications

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“…The mechanism and binding partners of the remaining eight RND proteins are as yet undefined (27). Of the four that are each located within a full complement operon, the cobalt-zinc-cadmium resistance proteins (CzcCBA) confer resistance to Cd(II), Zn(II), and Co(II), and the cobalt-nickel-resistance proteins (CnrCBA) confer resistance to Co(II) and Ni(II) (29)(30)(31)(32)(33)(34). zneBAC is also contained in a full complement operon, and we previously showed that the periplasmic MFP ZneB binds to Zn(II) with a K d of ∼3 μM (35), but not to Cd(II) or Co(II) (or other divalent metal ions).…”

Section: Resultsmentioning

confidence: 99%

Structures of intermediate transport states of ZneA, a Zn(II)/proton antiporter

Pak

Ekendé

Kifle

et al. 2013

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

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Efflux pumps belonging to the ubiquitous resistance-nodulationcell division (RND) superfamily transport substrates out of cells by coupling proton conduction across the membrane to a conformationally driven pumping cycle. The heavy metal-resistant bacteria Cupriavidus metallidurans CH34 relies notably on as many as 12 heavy metal efflux pumps of the RND superfamily. Here we show that C. metallidurans CH34 ZneA is a proton driven efflux pump specific for Zn(II), and that transport of substrates through the transmembrane domain may be electrogenic. We report two Xray crystal structures of ZneA in intermediate transport conformations, at 3.0 and 3.7 Å resolution. The trimeric ZneA structures capture protomer conformations that differ in the spatial arrangement and Zn(II) occupancies at a proximal and a distal substrate binding site. Structural comparison shows that transport of substrates through a tunnel that links the two binding sites, toward an exit portal, is mediated by the conformation of a short 14-aa loop. Taken together, the ZneA structures presented here provide mechanistic insights into the conformational changes required for substrate efflux by RND superfamily transporters.T he resistance-nodulation-cell division (RND) superfamily, named based on the original members' roles in metal resistance, root nodulation, and cell division (1), is found in all kingdoms of life and is comprised of nine phylogenetically distinct families (2-5). Functional characterization of RND proteins has shown that they are transmembrane efflux pumps that transport a variety of substrates out of cells, powered by an electrochemical proton gradient (6). In Gram-negative bacteria, RND pumps are specific for toxic substrates and largely belong to one of two families; the heavy metal efflux (HME) family and the multidrug hydrophobe/amphiphile efflux-1 (HAE1) family. For the HME-and HAE1-RND-driven efflux systems, a trimer of the RND pump in the plasma membrane is coupled by a hexamer of a periplasmic membrane fusion protein (MFP), also specific for the metal ion substrate in HME-RND-driven efflux systems, to a pore formed by a trimeric outer membrane factor (OMF), forming a continuous conduit that spans the inner and outer membranes (7-10). Each protomer of the RND trimer consists of a transmembrane domain of 12 transmembrane α-helices and two large hydrophilic loops that comprise the substratebinding porter (or pore) domain and the OMF-coupling docking domain (11).X-ray crystal structures of only four RND efflux pumps have been described, including just one of the HME family to date, shedding some light on the conformational changes that are necessary for transport (11)(12)(13)(14)(15)(16)(17)(18)(19)(20). In two of these structures, those of the HAE1 efflux pumps AcrB from Escherichia coli and of the closely related MexB from Pseudomonas aeruginosa, each protomer of the homotrimer is trapped in a unique conformation, with only a single protomer conducive for substrate binding, suggestive of a functionally rotating mechanism (12, ...

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Section: Resultsmentioning

confidence: 99%

Structures of intermediate transport states of ZneA, a Zn(II)/proton antiporter

Pak

Ekendé

Kifle

et al. 2013

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

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“…For example, iron in tomato apoplastic fluids was measured to be 2.2 to 4.2 M after collection by direct centrifugation (19). At these concentrations of iron, less-well-understood lower-affinity iron import systems are responsible for importing nutritional iron, potentially at a lower energetic cost (21,23). We show that DC3000 could readily utilize a number of different iron sources supplied at these levels (Fig.…”

mentioning

confidence: 78%

The Phytopathogen Pseudomonas syringae pv. tomato DC3000 Has Three High-Affinity Iron-Scavenging Systems Functional under Iron Limitation Conditions but Dispensable for Pathogenesis

Jones

Wildermuth

2011

J Bacteriol

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High-affinity iron scavenging through the use of siderophores is a well-established virulence determinant in mammalian pathogenesis. However, few examples have been reported for plant pathogens. Here, we use a genetic approach to investigate the role of siderophores in Pseudomonas syringae pv. tomato DC3000 (DC3000) virulence in tomato. DC3000, an agronomically important pathogen, has two known siderophores for highaffinity iron scavenging, yersiniabactin and pyoverdin, and we uncover a third siderophore, citrate, required for growth when iron is limiting. Though growth of a DC3000 triple mutant unable to either synthesize or import these siderophores is severely restricted in iron-limited culture, it is fully pathogenic. One explanation for this phenotype is that the DC3000 triple mutant is able to directly pirate plant iron compounds such as heme/hemin or iron-nicotianamine, and our data indicate that DC3000 can import iron-nicotianamine with high affinity. However, an alternative explanation, supported by data from others, is that the pathogenic environment of DC3000 (i.e., leaf apoplast) is not iron limited but is iron replete, with available iron of >1 M. Growth of the triple mutant in culture is restored to wild-type levels by supplementation with a variety of iron chelates at >1 M, including iron(III) dicitrate, a dominant chelate of the leaf apoplast. This suggests that lower-affinity iron import would be sufficient for DC3000 iron nutrition in planta and is in sharp contrast to the high-affinity iron-scavenging mechanisms required in mammalian pathogenesis.Pathogenic microorganisms must acquire all critical nutrients from the host environment in order to survive. Iron nutrition in particular presents a fundamental challenge due to its extremely low solubility in aerobic environments at moderate pHs (ϳ10 Ϫ9 M) and further host limitation of available free iron (e.g., ϳ10Ϫ18 M in mammalian fluids [22,44]). Most bacteria require much higher levels (i.e., 10 Ϫ6 to 10 Ϫ7 M of bioavailable iron) for optimal growth (24) and therefore must solve an iron supply problem for survival. There is abundant evidence that competition for iron is a key virulence determinant in mammalian pathosystems. Iron in human serum is predominantly complexed with transferrin protein (pFe ϭ 10 22 M [1]) and is therefore limited from invading pathogens. The human immune system can limit iron further through release of the iron-binding protein lactoferrin at the site of infection (57). Evading these defenses, several mammalian pathogens pirate host-synthesized heme for iron nutrition directly from host fluids (20). Alternatively, pathogenic bacteria can scavenge iron from host fluids using very high-affinity iron carriers termed siderophores that can effectively compete with host iron chelates for iron. Siderophores are low-molecular-weight compounds that are synthesized, exported from the bacterial cell, and then imported once bound to iron (43). As this strategy carries metabolic costs associated with siderophore synthesis and transp...

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“…ZraP is a metal-binding protein that is induced by a two-component system that responds to zinc or lead (33,42), while CnrX is thought to act as a metal sensor that binds cobalt, nickel, and potentially copper in an extracytoplasmic function (ECF)-sigma factor signaling pathway (18,19,34,38,61). Interestingly, copper ions have been implicated as an inducer of the Cpx pathway (26,(67)(68)(69).…”

Section: Discussionmentioning

confidence: 99%

Structure of the Periplasmic Stress Response Protein CpxP

et al. 2011

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CpxP is a novel bacterial periplasmic protein with no homologues of known function. In Gram-negative enteric bacteria, CpxP is thought to interact with the two-component sensor kinase, CpxA, to inhibit induction of the Cpx envelope stress response in the absence of protein misfolding. CpxP has also been shown to facilitate DegP-mediated proteolysis of misfolded proteins. Six mutations that negate the ability of CpxP to function as a signaling protein are localized in or near two conserved LTXXQ motifs that define a class of proteins with similarity to CpxP, Pfam PF07813. To gain insight into how these mutations might affect CpxP signaling and/or proteolytic adaptor functions, the crystal structure of CpxP from Escherichia coli was determined to 2.85-Å resolution. The structure revealed an antiparallel dimer of intertwined ␣-helices with a highly basic concave surface. Each protomer consists of a long, hooked and bent hairpin fold, with the conserved LTXXQ motifs forming two diverging turns at one end. Biochemical studies demonstrated that CpxP maintains a dimeric state but may undergo a slight structural adjustment in response to the inducing cue, alkaline pH. Three of the six previously characterized cpxP loss-of-function mutations, M59T, Q55P, and Q128H, likely result from a destabilization of the protein fold, whereas the R60Q, D61E, and D61V mutations may alter intermolecular interactions.

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Control of Expression of a Periplasmic Nickel Efflux Pump by Periplasmic Nickel Concentrations

Cited by 64 publications

References 52 publications

Structures of intermediate transport states of ZneA, a Zn(II)/proton antiporter

Structures of intermediate transport states of ZneA, a Zn(II)/proton antiporter

The Phytopathogen Pseudomonas syringae pv. tomato DC3000 Has Three High-Affinity Iron-Scavenging Systems Functional under Iron Limitation Conditions but Dispensable for Pathogenesis

Structure of the Periplasmic Stress Response Protein CpxP

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