Conserved residues modulate copper release in human copper chaperone Atox1

It is unclear how the human copper (Cu) chaperone Atox1 delivers Cu to metal-binding domains of Wilson and Menkes disease proteins in the cytoplasm. To begin to address this problem, we have characterized Cu(I) release from wild-type Atox1 and two point mutants (Met₁₀Ala and Lys₆₀Ala). The dynamics...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2008-08, Vol.105 (32), p.11158-11163
Main Authors: Hussain, Faiza, Olson, John S, Wittung-Stafshede, Pernilla
Format: Article
Language:English
Subjects:
Adenosine triphosphatases
Adenosine Triphosphatases - chemistry
Adenosine Triphosphatases - genetics
Adenosine Triphosphatases - metabolism
Amino acids
Animals
Binding sites
Biochemistry
Biological Sciences
Cation Transport Proteins - chemistry
Cation Transport Proteins - genetics
Cation Transport Proteins - metabolism
Cattle
Chelating Agents - chemistry
Copper
Copper - chemistry
Copper - metabolism
Copper-transporting ATPases
Hepatolenticular degeneration
Humans
Kinetics
Ligands
Menkes kinky hair syndrome
Metallochaperones
Models, Chemical
Molecular Chaperones - chemistry
Molecular Chaperones - genetics
Molecular Chaperones - metabolism
Molecules
Multiprotein Complexes - chemistry
Multiprotein Complexes - genetics
Multiprotein Complexes - metabolism
Mutation
Point Mutation
Protein Binding - physiology
Proteins
Quinolines - chemistry
Serum Albumin, Bovine - chemistry
Serum Albumin, Bovine - genetics
Serum Albumin, Bovine - metabolism
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Summary:It is unclear how the human copper (Cu) chaperone Atox1 delivers Cu to metal-binding domains of Wilson and Menkes disease proteins in the cytoplasm. To begin to address this problem, we have characterized Cu(I) release from wild-type Atox1 and two point mutants (Met₁₀Ala and Lys₆₀Ala). The dynamics of Cu(I) displacement from holo-Atox1 were measured by using the Cu(I) chelator bicinchonic acid (BCA) as a metal acceptor. BCA removes Cu(I) from Atox1 in a three-step process involving the bimolecular formation of an initial Atox1-Cu-BCA complex followed by dissociation of Atox1 and the binding of a second BCA to generate apo-Atox1 and Cu-BCA₂. Both mutants lose Cu(I) more readily than wild-type Atox1 because of more rapid and facile displacement of the protein from the Atox1-Cu-BCA intermediate by the second BCA. Remarkably, Cu(I) uptake from solution by BCA is much slower than the transfer from holo-Atox1, presumably because of slow dissociation of DTT-Cu complexes. These results suggest that Cu chaperones play a key role in making Cu(I) rapidly accessible to substrates and that the activated protein-metal-chelator complex may kinetically mimic the ternary chaperone-metal-target complex involved in Cu(I) transfer in vivo.
ISSN:0027-8424
1091-6490
1091-6490
DOI:10.1073/pnas.0802928105