Mechanism of CuA assembly

Abriata, Luciano A.; Banci, Lucia; Bertini, Ivano; Ciofi‐Baffoni, Simone; Gkazonis, Petros; Spyroulias, Georgios A.; Vila, Alejandro J.; Wang, Shenlin

doi:10.1038/nchembio.110

Cited by 114 publications

(206 citation statements)

References 23 publications

(15 reference statements)

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“…We have recently elucidated the chaperone-mediated mechanism of copper uptake by the Cu A -binding cupredoxin domain in Thermus thermophilus ba 3 cytochrome c oxidase (41). In that work, we observed that the NMR spectra of the apo and metal-lated forms of this protein show significant differences, suggesting that there might be nonnegligible structural perturbations upon metal binding, consistent with the finding that copper uptake is mediated by a metallochaperone (Fig.…”

supporting

confidence: 75%

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Flexibility of the metal-binding region in apo-cupredoxins

Zaballa

Abriata

Donaire

et al. 2012

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

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Protein-mediated electron transfer is an essential event in many biochemical processes. Efficient electron transfer requires the reorganization energy of the redox event to be minimized, which is ensured by the presence of rigid donor and acceptor sites. Electron transfer copper sites are present in the ubiquitous cupredoxin fold, able to bind one or two copper ions. The low reorganization energy in these metal centers has been accounted for by assuming that the protein scaffold creates an entatic/rack-induced state, which gives rise to a rigid environment by means of a preformed metal chelating site. However, this notion is incompatible with the need for an exposed metal-binding site and protein-protein interactions enabling metallochaperone-mediated assembly of the copper site. Here we report an NMR study that reveals a high degree of structural heterogeneity in the metal-binding region of the nonmetallated Cu A -binding cupredoxin domain, arising from microsecond to second dynamics that are quenched upon metal binding. We also report similar dynamic features in apo-azurin, a paradigmatic blue copper protein, suggesting a general behavior. These findings reveal that the entatic/rack-induced state, governing the features of the metal center in the copper-loaded protein, does not require a preformed metal-binding site. Instead, metal binding is a major contributor to the rigidity of electron transfer copper centers. These results reconcile the seemingly contradictory requirements of a rigid, occluded center for electron transfer, and an accessible, dynamic site required for in vivo copper uptake.metalloproteins | nuclear magnetic resonance | protein dynamics

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supporting

confidence: 75%

“…The copper cargo in the Cu A -binding cupredoxin domain of T. thermophilus oxidase is due to the action of a specific Cu(I)-metallochaperone (PCu A C) (41). Similar copper-loading mechanisms have been proposed for the type 1 protein plastocyanin (63).…”

Section: Discussionmentioning

confidence: 73%

Flexibility of the metal-binding region in apo-cupredoxins

Zaballa

Abriata

Donaire

et al. 2012

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

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“…Thus, the apo form of a newly synthesized COX II is expected to have the two Cys ligands at least partially oxidized, being therefore unable to bind the Cu(I) ions. Sco proteins have been suggested to act as thiol reductases of the Cys ligands of COX II based on their thioredoxin fold (27,37), which is indeed supported by in vitro experiments on the bacterial thermophilic Sco protein (32,37). Based on the reduction potentials of the S-S/2SH redox couple of the human proteins: Cox17 (−198 mV) > Sco1 (−277 mV) > COX II* (−288 mV) > Sco2 (less than −300 mV) (24,38,39), Sco2 would be the only protein able to reduce the disulfide bond in COX II*.…”

Section: Copper-bound Human Sco2 Is a Thiol Oxidoreductase Of The Cysmentioning

confidence: 77%

“…Removal of this transmembrane stretch has allowed the expression of soluble COX II domains from several bacterial oxidases (30,31). Among them, the soluble fragment from Thermus thermophilus COX II (Tt COX II hereafter) is the only one stable for long periods of time and under different conditions (32,33). Instead, no eukaryotic COX II subunit has been expressed in a stable, soluble form to date.…”

Section: Resultsmentioning

confidence: 99%

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Loop recognition and copper-mediated disulfide reduction underpin metal site assembly of Cu _A in human cytochrome oxidase

Morgada

Abriata

Cefaro

et al. 2015

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

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Maturation of cytochrome oxidases is a complex process requiring assembly of several subunits and adequate uptake of the metal cofactors. Two orthologous Sco proteins (Sco1 and Sco2) are essential for the correct assembly of the dicopper Cu A site in the human oxidase, but their function is not fully understood. Here, we report an in vitro biochemical study that shows that Sco1 is a metallochaperone that selectively transfers Cu(I) ions based on loop recognition, whereas Sco2 is a copper-dependent thiol reductase of the cysteine ligands in the oxidase. Copper binding to Sco2 is essential to elicit its redox function and as a guardian of the reduced state of its own cysteine residues in the oxidizing environment of the mitochondrial intermembrane space (IMS). These results provide a detailed molecular mechanism for Cu A assembly, suggesting that copper and redox homeostasis are intimately linked in the mitochondrion.

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Biochemical pathway for the biosynthesis of the Cu_A center in bacterial cytochrome c oxidase

2019

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The di‐copper center CuA is an essential metal cofactor in cytochrome oxidase (Cox) of mitochondria and many prokaryotes, mediating one‐electron transfer from cytochrome c to the site for oxygen reduction. CuA is located in subunit II (CoxB) of Cox and protrudes into the periplasm of Gram‐negative bacteria or the mitochondrial intermembrane space. How the two copper ions are brought together to build CoxB·CuA is the subject of this review. It had been known that the reductase TlpA and the metallochaperones ScoI and PcuC are required for CuA formation in bacteria, but the mechanism of copper transfer has emerged only recently for the Bradyrhizobium diazoefficiens system. It consists of the following steps: (a) TlpA keeps the active site cysteine pair of CoxB in its dithiol state as a prerequisite for metal insertion; (b) ScoI·Cu2+ rapidly forms a transient complex with apo‐CoxB; (c) PcuC, loaded with Cu1+ and Cu2+, dissociates this complex to CoxB·Cu2+, and a second PcuC·Cu1+·Cu2+ transfers Cu1+ to CoxB·Cu2+, yielding mature CoxB·CuA. Variants of this pathway might exist in other bacteria or mitochondria.

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Mechanism of CuA assembly

Cited by 114 publications

References 23 publications

Flexibility of the metal-binding region in apo-cupredoxins

Flexibility of the metal-binding region in apo-cupredoxins

Loop recognition and copper-mediated disulfide reduction underpin metal site assembly of Cu _A in human cytochrome oxidase

Biochemical pathway for the biosynthesis of the Cu_A center in bacterial cytochrome c oxidase

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Mechanism of CuA assembly

Cited by 114 publications

References 23 publications

Flexibility of the metal-binding region in apo-cupredoxins

Flexibility of the metal-binding region in apo-cupredoxins

Loop recognition and copper-mediated disulfide reduction underpin metal site assembly of Cu A in human cytochrome oxidase

Biochemical pathway for the biosynthesis of the CuA center in bacterial cytochrome c oxidase

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Resources

About

Loop recognition and copper-mediated disulfide reduction underpin metal site assembly of Cu _A in human cytochrome oxidase

Biochemical pathway for the biosynthesis of the Cu_A center in bacterial cytochrome c oxidase