Structure and Metal Loading of a Soluble Periplasm Cuproprotein

Waldron, Kevin J.; Firbank, S.J.; Dainty, Samantha J.; Pérez-Rama, Mónica; Tottey, Steve; Robinson, Nigel J.

doi:10.1074/jbc.m110.153080

Cited by 31 publications

(24 citation statements)

References 39 publications

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“…1 A and B). This multiprotein complex is not found in periplasm extracts (28), and is also present in the related cyanobacterium Synechococcus PCC 7942 (SI Appendix). Cu-P1 is a cytoplasmic protein complex which obtains copper from a route which is independent of both ATPases, and moreover CtaA, but not PacS, competes with Cu-P1 for copper ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Cyanobacterial metallochaperone inhibits deleterious side reactions of copper

Tottey

Patterson

Banci

et al. 2011

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

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115

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Copper metallochaperones supply copper to cupro-proteins through copper-mediated protein-protein-interactions and it has been hypothesized that metallochaperones thereby inhibit copper from causing damage en route. Evidence is presented in support of this latter role for cyanobacterial metallochaperone, Atx1. In cyanobacteria Atx1 contributes towards the supply of copper to plastocyanin inside thylakoids but it is shown here that in copper-replete medium, copper can reach plastocyanin without Atx1. Unlike metallochaperone- independent copper-supply to superoxide dismutase in eukaryotes, glutathione is not essential for Atx1- independent supply to plastocyanin: Double mutants missing atx1 and gshB (encoding glutathione synthetase) accumulate the same number of atoms of copper per cell in the plastocyanin pool as wild type. Critically, atx1gshB are hypersensitive to elevated copper relative to wild type cells and also relative to gshB single mutants with evidence that hyp ersensitivity arises due to the mislocation of copper to sites for other metals including iron and zinc. The zinc site on the amino-terminal domain (ZiaA N) of the P 1-type zinctransporting ATPase is especially similar to the copper site of the Atx1 target PacS N, and ZiaA N will bind Cu(I) more tightly than zinc. An NMR model of a substituted-ZiaA N-Cu(I)-Atx1 heterodimer has been generated making it possible to visualize a juxtaposition of residues surrounding the ZiaA N zinc site, including Asp 18, which normally repulse Atx1. Equivalent repulsion between bacterial copper metallochaperones and the amino-terminal regions of P 1-type ATPases for metals other than Cu(I) is conserved, again consistent with a role for copper metallochaperones to withhold copper from binding sites for other metals

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Section: Resultsmentioning

confidence: 99%

Cyanobacterial metallochaperone inhibits deleterious side reactions of copper

Tottey

Patterson

Banci

et al. 2011

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

Self Cite

115

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“…The extracellular release pathway is possibly involved in specific and controlled delivery to Cu + acceptor chaperones 30 , preventing detrimental effects of free copper in the extracellular space, or to other cupro-proteins that need the ion for function 31,32 . The pathway further explains why Menkes and Wilson mutations in this region impair ATP7A/B function.…”

Section: Discussionmentioning

confidence: 99%

Copper-transporting P-type ATPases use a unique ion-release pathway

Andersson

Mattle

Sitsel

et al. 2013

Nat Struct Mol Biol

103

156

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Heavy metals in cells are typically regulated by PIB-type ATPases such as the copper transporting Cu+-ATPases. The first crystal structure of a Cu+-ATPase (LpCopA) was trapped in a transition state of dephosphorylation (E2.Pi) and inferred to be occluded. The structure revealed a PIB-specific topology and suggested a copper transport pathway across the membrane. Here we show by molecular dynamics (MD) simulations that extracellular water solvates the transmembrane (TM) domain, indicative of a pathway for Cu+ release. Furthermore, a new LpCopA crystal structure determined at 2.8 Å resolution, trapped in the E2P state (which is associated with extracellular exchange in PII-type ATPases), delineates the same conduit as also further supported by site-directed mutagenesis. The E2P and E2.Pi states therefore appear equivalent and open to the extracellular side, in contrast to PII-type ATPases where the E2.Pi state is occluded. This indicates that Cu+-ATPases couple dephosphorylation differently to the conformational changes associated with ion extrusion. The ion pathway may explain why Menkes’ and Wilson’s disease mutations at the extracellular side impair protein function, and points to an accessible site for novel inhibitors targeting Cu+-ATPases of pathogens.

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“…Limited information about copper acquisition within cellular compartments is currently available, but appears to involve complex processes in both prokaryotes and eukaryotes (36,66). The presence and nature of available copper pools in the thylakoid are unknown, but tight binding of Cu(I) by the transmembrane sites of the copper transporters would make Cu(I) release to solution unlikely.…”

Section: Discussionmentioning

confidence: 99%

“…CucA (a quercetin dioxygenase), the most abundant soluble periplasmic copper protein in Synechocystis is exported unfolded and metal-free to this compartment preventing the need for more labile cytosolic copper pools which could result in the mis-metallation of metalloproteins (11). However, CucA does not directly acquire copper from these periplasmic pools in vivo (36), and CtaA, PacS N , and Atx1 have all been shown to be involved in copper loading CucA, although the mechanism of this process is unknown. The lack of a copper affinity gradient in the cytoplasm of Synechocystis (Fig.…”

Section: Discussionmentioning

confidence: 99%

Thermodynamics of copper and zinc distribution in the cyanobacterium Synechocystis PCC 6803

Badarau

Dennison

2011

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

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Copper is supplied to plastocyanin for photosynthesis and cytochrome c oxidase for respiration in the thylakoids of Synechocystis PCC 6803 by the membrane-bound P-type ATPases CtaA and PacS, and the metallochaperone Atx1. We have determined the Cu(I) affinities of all of the soluble proteins and domains in this pathway. The Cu(I) affinities of the trafficking proteins range from 5 × 10 16 to 5 × 10 17 M -1 at pH 7.0, consistent with values for homologues. Unusually, Atx1 binds Cu(I) significantly tighter than the metal-binding domains (MBDs) of CtaA and PacS (CtaA N and PacS N ), and equilibrium copper exchange constants of approximately 0.2 are obtained for transfer to the MBDs. Dimerization of Atx1 increases the affinity for Cu(I), but the loop 5 His61 residue has little influence. The MBD of the zinc exporter ZiaA (ZiaA N ) exhibits an almost identical Cu(I) affinity, and Cu(I) exchange with Atx1, as CtaA N and PacS N , and the relative stabilities of the complexes must enable the metallochaperone to distinguish between the MBDs. The binding of potentially competing zinc to the trafficking proteins has been studied. ZiaA N has the highest Zn(II) affinity and thermodynamics could be important for zinc removal from the cell. Plastocyanin has a Cu(I) affinity of 2.6 × 10 17 M -1 , 15-fold tighter than that of the Cu A site of cytochrome c oxidase, highlighting the need for specific mechanisms to ensure copper delivery to both of these targets. The narrow range of Cu(I) affinities for the cytoplasmic copper proteins in Synechocystis will facilitate relocation when copper is limiting.

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Structure and Metal Loading of a Soluble Periplasm Cuproprotein

Cited by 31 publications

References 39 publications

Cyanobacterial metallochaperone inhibits deleterious side reactions of copper

Cyanobacterial metallochaperone inhibits deleterious side reactions of copper

Copper-transporting P-type ATPases use a unique ion-release pathway

Thermodynamics of copper and zinc distribution in the cyanobacterium Synechocystis PCC 6803

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