Copper chaperone Atox1 interacts with the metal-binding domain of Wilson's disease protein in cisplatin detoxification

Human copper transporters ATP7B (Wilson's disease protein) and ATP7A (Menkes' disease protein) have been implicated in tumour resistance to cisplatin, a widely used anticancer drug. Cisplatin binds to the copper-binding sites in the N-terminal domain of ATP7B, and this binding may be an essential step of cisplatin detoxification involving copper ATPases. In the present study, we demonstrate that cisplatin and a related platinum drug carboplatin produce the same adduct following reaction with MBD2 [metal-binding domain (repeat) 2], where platinum is bound to the side chains of the cysteine residues in the CxxC copper-binding motif. This suggests the same mechanism for detoxification of both drugs by ATP7B. Platinum can also be transferred to MBD2 from copper chaperone Atox1, which was shown previously to bind cisplatin. Binding of the free cisplatin and reaction with the cisplatin-loaded Atox1 produce the same protein-bound platinum intermediate. Transfer of platinum along the copper-transport pathways in the cell may serve as a mechanism of drug delivery to its target in the cell nucleus, and explain tumour-cell resistance to cisplatin associated with the overexpression of copper transporters ATP7B and ATP7A.

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Molecular modelling of the nucleotide-binding domain of Wilson's disease protein: location of the ATP-binding site, domain dynamics and potential effects of the major disease mutations

Biochemical Journal ◽

10.1042/bj20040326 ◽

2004 ◽

Vol 382 (1) ◽

pp. 293-305 ◽

Cited By ~ 16

Author(s):

Roman G. EFREMOV ◽

Yuri A. KOSINSKY ◽

Dmitry E. NOLDE ◽

Ruslan TSIVKOVSKII ◽

Alexander S. ARSENIEV ◽

...

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Molecular Modelling ◽

Wilson’S Disease ◽

Wilson's Disease ◽

Binding Domain ◽

Atp Binding ◽

Disease Mutations ◽

Disease Protein ◽

Dynamics Simulations

WNDP (Wilson's disease protein) is a copper-transporting ATPase that plays an essential role in human physiology. Mutations in WNDP result in copper accumulation in tissues and cause a severe hepato-neurological disorder known as Wilson's disease. Several mutations were surmised to affect the nucleotide binding and hydrolysis by WNDP; however, how the nucleotides bind to normal and mutated WNDP remains unknown. To aid such studies, we performed the molecular modelling of the spatial structure and dynamics of the ATP-binding domain of WNDP and its interactions with ATP. The three-dimensional models of this domain in two conformations were built using the X-ray structures of the Ca2+-ATPase in the E1 and E2 states. To study the functional aspects of the models, they were subjected to long-term molecular dynamics simulations in an explicit solvent; similar calculations were performed for the ATP-binding domain of Ca2+-ATPase. In both cases, we found large-scale motions that lead to significant changes of distances between several functionally important residues. The ATP docking revealed two possible modes of ATP binding: via adenosine buried in the cleft near residues H1069, R1151 and D1164, and via phosphate moiety ‘anchored’ by H-bonds with residues in the vicinity of catalytic D1027. Furthermore, interaction of ATP with both sites occurs if they are spatially close to each other. This may be achieved after relative domain motions of the ‘closure’ type observed in molecular dynamics simulations. The results provide a framework for analysis of disease mutations and for future mutagenesis studies.

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Expression, purification and copper-binding studies of the first metal-binding domain of Menkes protein

European Journal of Biochemistry ◽

10.1046/j.1432-1327.1999.00680.x ◽

1999 ◽

Vol 264 (3) ◽

pp. 890-896 ◽

Cited By ~ 14

Author(s):

Pia Y. Jensen ◽

Nicklas Bonander ◽

Nina Horn ◽

Zeynep Tumer ◽

Ole Farver

Keyword(s):

Metal Binding ◽

Binding Domain ◽

Copper Binding ◽

Binding Studies ◽

Metal Binding Domain ◽

Menkes Protein

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Copper-mediated dimerization of CopZ, a predicted copper chaperone from Bacillus subtilis

Biochemical Journal ◽

10.1042/bj20021036 ◽

2002 ◽

Vol 368 (3) ◽

pp. 729-739 ◽

Cited By ~ 45

Author(s):

Margaret A. KIHLKEN ◽

Andrew P. LEECH ◽

Nick E. LE BRUN

Keyword(s):

Bacillus Subtilis ◽

Metal Binding ◽

Metal Ion ◽

Complex Nature ◽

Binding Motif ◽

Copper Chaperone ◽

Ion Transfer ◽

Copper Binding ◽

Binding Properties ◽

In The Wild

Understanding the metal-binding properties and solution states of metallo-chaperones is a key step in understanding how they function in metal ion transfer. Using spectroscopic, bioanalytical and biochemical methods, we have investigated the copper-binding properties and association states of the putative copper chaperone of Bacillus subtilis, CopZ, and a variant of the protein lacking the two cysteine residues of the MXCXXC copper-binding motif. We show that copper-free CopZ exists as a monomer, but that addition of copper(I) causes the protein to associate into homodimers. The nature of the copper(I)—CopZ complex is dependent on the level of copper loading, and we report the detection of three distinct forms, containing 0.5, 1.0 and 1.5 copper(I) ions per protein. The presence of excess dithiothreitol has a significant effect on copper(I) binding to CopZ, such that, in its presence, copper(I)—CopZ occurs mainly as a monomer species. Data for copper binding to the double-cysteine variant of CopZ are consistent with an essential role for these residues in tight copper binding in the wild-type protein. We conclude that the complex nature of copper(I) binding to CopZ may underpin mechanisms of protein-to-protein copper(I) transfer.

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Copper-Transfer Mechanism from the Human Chaperone Atox1 to a Metal-Binding Domain of Wilson Disease Protein

The Journal of Physical Chemistry B ◽

10.1021/jp911208z ◽

2010 ◽

Vol 114 (10) ◽

pp. 3698-3706 ◽

Cited By ~ 36

Author(s):

Agustina Rodriguez-Granillo ◽

Alejandro Crespo ◽

Dario A. Estrin ◽

Pernilla Wittung-Stafshede

Keyword(s):

Metal Binding ◽

Wilson Disease ◽

Binding Domain ◽

Transfer Mechanism ◽

Wilson Disease Protein ◽

Metal Binding Domain ◽

Disease Protein ◽

Copper Transfer

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Copper–zinc superoxide dismutase (Sod1) activation terminates interaction between its copper chaperone (Ccs) and the cytosolic metal-binding domain of the copper importer Ctr1

BioMetals ◽

10.1007/s10534-019-00206-3 ◽

2019 ◽

Vol 32 (4) ◽

pp. 695-705 ◽

Cited By ~ 7

Author(s):

Amélie Skopp ◽

Stefanie D. Boyd ◽

Morgan S. Ullrich ◽

Li Liu ◽

Duane D. Winkler

Keyword(s):

Superoxide Dismutase ◽

Metal Binding ◽

Binding Domain ◽

Copper Chaperone ◽

Copper Zinc Superoxide Dismutase ◽

Zinc Superoxide Dismutase ◽

Metal Binding Domain ◽

Copper Zinc

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The Distinct Roles of the N-terminal Copper-binding Sites in Regulation of Catalytic Activity of the Wilson's Disease Protein

Journal of Biological Chemistry ◽

10.1074/jbc.m305408200 ◽

2003 ◽

Vol 278 (34) ◽

pp. 32212-32218 ◽

Cited By ~ 79

Author(s):

Dominik Huster ◽

Svetlana Lutsenko

Keyword(s):

Catalytic Activity ◽

Binding Sites ◽

Wilson’S Disease ◽

Wilson's Disease ◽

Copper Binding ◽

Disease Protein ◽

Copper Binding Sites

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The soluble metal-binding domain of the copper transporter ATP7B binds and detoxifies cisplatin

Biochemical Journal ◽

10.1042/bj20081359 ◽

2009 ◽

Vol 419 (1) ◽

pp. 51-59 ◽

Cited By ~ 46

Author(s):

Nataliya V. Dolgova ◽

Doug Olson ◽

Svetlana Lutsenko ◽

Oleg Y. Dmitriev

Keyword(s):

Metal Binding ◽

Binding Sites ◽

Active Drug ◽

Binding Domain ◽

Copper Binding ◽

Protein Fragment ◽

Copper Transporter ◽

Metal Binding Domain ◽

Cell Expression

Wilson disease ATPase (ATP7B) has been implicated in the resistance of cancer cells to cisplatin. Using a simple in vivo assay in bacterial culture, in the present study we demonstrate that ATP7B can confer resistance to cisplatin by sequestering the drug in its N-terminal metal-binding domain without active drug extrusion from the cell. Expression of a protein fragment containing four N-terminal MBRs (metal-binding repeats) of ATP7B (MBR1–4) protects cells from the toxic effects of cisplatin. One MBR1–4 molecule binds up to three cisplatin molecules at the copper-binding sites in the MBRs. The findings of the present study suggest that suppressing enzymatic activity of ATP7B may not be an effective way of combating cisplatin resistance. Rather, the efforts should be directed at preventing cisplatin binding to the protein.

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