Characterisation of copper-binding to the second sub-domain of the Menkes protein ATPase (MNKr2).
The Menkes ATPase (MNK) has an essential role in the translocation of copper across cellular membranes. In a complementary manner, the intracellular concentration of copper regulates the activity and cellular location of the ATPase through its six homologous amino-terminal domains. The roles of the six amino-terminal domains in the activation and cellular trafficking processes are unknown. Understanding the role of these domains relies on the development of an understanding of their metal-binding properties and structural properties. The second conserved sub-domain of MNK was over-expressed, purified and its copper-binding properties characterised. Reconstitution studies demonstrate that copper binds to MNKr2 as Cu(I) with a stoichiometry of one copper per domain. This is the first direct evidence of copper-binding to the MNK amino-terminal repeats. Circular dichroism studies suggest that the binding or loss of copper to MNKr2 does not cause substantial changes to the secondary structure of the protein.
Harrison
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MD
National Research Centre for Environmental Toxicology, The University of Queensland, Coopers Plains 4108, Australia.
Meier
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S
Dameron
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CT
D000251:Adenosine Triphosphatases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D002352:Carrier Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D027682:Cation Transport Proteins; D002942:Circular Dichroism; D003300:Copper / Q000737:chemistry*; D000073840:Copper-Transporting ATPases; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013056:Spectrophotometry, Ultraviolet
D000596:Amino Acids; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D011993:Recombinant Fusion Proteins; D003300:Copper; D000251:Adenosine Triphosphatases; C081184:ATP7A protein, human; D000073840:Copper-Transporting ATPases