Solution Structure of the Apo and Copper(I)-Loaded Human Metallochaperone HAH1

Anastassopoulou, Ioanna; Banci, Lucia; Bertini, Ivano; Cantini, Francesca; Katsari, Efthalia; Rosato, Antonio

doi:10.1021/bi0487591

Cited by 121 publications

(173 citation statements)

References 45 publications

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“…[Runs were performed and analyzed for us by B. Demeler and A. Musatov (University of Texas Health Science Center, San Antonio, TX).] Previous NMR data also shows holo-Atox1 as a monomer in solution (37).…”

Section: Discussionmentioning

confidence: 73%

Conserved residues modulate copper release in human copper chaperone Atox1

Hussain¹,

Olson

Wittung-Stafshede

2008

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

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Section: Discussionmentioning

confidence: 73%

Conserved residues modulate copper release in human copper chaperone Atox1

Hussain¹,

Olson

Wittung-Stafshede

2008

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

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“…Sequence alignments and homology models on the six metal-binding domains of WLNP demonstrate that the Ϸ70 aa domains of N-WLNP are likely to be folded very similarly into ferredoxin-like units (16). This same fold is also found in both NMR and x-ray structures of the yeast Atx1 and human HAH1 metallochaperones (17)(18)(19)(20), the soluble cytosolic proteins that deliver copper to the copper-transporting P-type ATPases (21).…”

mentioning

confidence: 71%

Structure of human Wilson protein domains 5 and 6 and their interplay with domain 4 and the copper chaperone HAH1 in copper uptake

Achila

Banci

Bertini

et al. 2006

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

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146

252

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Human Wilson protein is a copper-transporting ATPase located in the secretory pathway possessing six N-terminal metal-binding domains. Here we focus on the function of the metal-binding domains closest to the vesicular portion of the copper pump, i.e., domain 4 (WLN4), and a construct of domains 5 and 6 (WLN5-6). For comparison purposes, some experiments were also performed with domain 2 (WLN2). The solution structure of apoWLN5-6 consists of two ferredoxin folds connected by a short linker, and 15 N relaxation rate measurements show that it behaves as a unit in solution. An NMR titration of apoWLN5-6 with the metallochaperone Cu(I)HAH1 reveals no complex formation and no copper exchange between the two proteins, whereas titration of Cu(I)HAH1 with WLN4 shows the formation of an adduct that is in fast exchange on the NMR time scale with the isolated protein species as confirmed by 15 N relaxation data. A similar interaction is also observed between Cu(I)HAH1 and WLN2; however, the relative amount of the adduct in the protein mixture is lower. An NMR titration of apoWLN5-6 with Cu(I)WLN4 shows copper transfer, first to WLN6 then to WLN5, without the formation of an adduct. Therefore, we suggest that WLN4 and WLN2 are two acceptors of Cu(I) from HAH1, which then somehow route copper to WLN5-6, before the ATP-driven transport of copper across the vesicular membrane.ATPase function ͉ copper transport ͉ metal-binding domain ͉ metallochaperone ͉ Wilson disease protein

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“…HAH1 samples were prepared as already reported, always without a poly-His tag (38). In titration experiments, we added copper(I)-HAH1 and apo-WLN1-6 directly in the NMR tube under N 2 atmosphere, using the same procedure previously followed for MNK1-6 (28).…”

Section: Methodsmentioning

confidence: 99%

An NMR Study of the Interaction of the N-terminal Cytoplasmic Tail of the Wilson Disease Protein with Copper(I)-HAH1

Banci

Bertini

Cantini

et al. 2009

Journal of Biological Chemistry

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118

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ATP7B is a human P 1B -type ATPase that has a crucial role in maintaining copper(I) homeostasis. Mutations in the corresponding gene are the cause of Wilson disease. Among its various distinguishing features is a long (ϳ630 amino acids) N-terminal cytosolic tail containing six domains that are individually folded and capable of binding one copper(I) ion each. We expressed the entire tail as a single construct in Escherichia coli and investigated its interaction with its copper chaperone (i.e. HAH1) by solution NMR spectroscopy. We observed that all six of the metal-binding domains were metallated by Cu(I)-HAH1, with the first, the second, and the fourth domains forming an adduct with it. This behavior is different from that of the highly similar human ATPase ATP7A, in which only two domains form such an adduct. The distinct behaviors of the different domains were analyzed in terms of the energetics of Cu(I) transfer, hinting at a specific role of the interaction with copper(I)-HAH1 in the overall functional process.Human ATP7B is a P 1B -type ATPase that, like the other human copper(I)-transporting ATPase, ATP7A, can translocate copper(I) to the trans-Golgi network, where the metal is incorporated into copper-dependent enzymes. Copper stimulation results in the redistribution of both ATP7A and ATP7B to the plasma membrane through intracellular vesicles in order to export excess copper out of the cytosol (1-4). Mutations in the ATP7B autosomal gene produce the copper accumulation in tissues that characterizes Wilson disease (5-7). Mutations in ATP7A are instead responsible for Menkes disease (8, 9). ATP7A and ATP7B are often referred to as the MNK and WLN proteins, respectively. P 1B -type ATPases contain four major regions/domains (the N-terminal copper-binding tail, the transmembrane domain, the ATP-binding domain, and the phosphatase domain) as well as a short C-terminal tail (10). The N-terminal copper-binding cytosolic tails of WLN and MNK are both ϳ630 amino acids long. They contain six 70-amino acid, independently folded metal-binding domains (MBDs), 2 which are relatively similar in sequence and structure (11)(12)(13)(14)(15). Each MBD harbors the conserved sequence motif GMXCXXC, through which it can bind one equivalent of copper(I) (16). The N-terminal tail has an important role in tuning the activity of the enzyme (17-19) and in modulating the intracellular trafficking rates of ATP7A and ATP7B (20 -22), an aspect of the cellular function of these ATPases that is lacking in less complex organisms, including yeast. The presence of either the intact fifth or intact sixth metal-binding domain in the human proteins is sufficient to support both WLN/MNK activity and intracellular trafficking at essentially normal levels (21,(23)(24)(25)(26). This has stimulated researchers to investigate the cellular role of the additional four domains that are present in the two human ATPases as well as in several other mammalian homologues. The K Cu binding affinity of the six MBDs of both the MNK (13, 27, 28) and WLN (29...

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Solution Structure of the Apo and Copper(I)-Loaded Human Metallochaperone HAH1

Cited by 121 publications

References 45 publications

Conserved residues modulate copper release in human copper chaperone Atox1

Conserved residues modulate copper release in human copper chaperone Atox1

Structure of human Wilson protein domains 5 and 6 and their interplay with domain 4 and the copper chaperone HAH1 in copper uptake

An NMR Study of the Interaction of the N-terminal Cytoplasmic Tail of the Wilson Disease Protein with Copper(I)-HAH1

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