Protein Interactions that Enable Safe and Efficient Copper Ion Transport in the Human Cytoplasm
Journal article, 2016

Although copper (Cu) is an essential metal for most living organisms, high levels and free such ions are toxic. In humans, after cellular uptake via the membrane-bound importer Ctr1, Cu is transported to targets by cytoplasmic Cu chaperones: Atox1 delivers Cu to membrane-bound P1B-type ATPases (i.e., ATP7B or Wilson disease protein) in the Golgi (secretory path; here, most Cu-dependent enzymes are loaded with Cu) whereas CCS delivers Cu specifically to cytoplasmic superoxide dismutase. In contrast to bacterial andyeast homologs, ATP7B has six similar metal-binding domains protruding into the cytoplasm: possibly, conformational changes among these regulate overall ATP7B activity. To reveal underlying molecular mechanisms as well as thermodynamic and kinetic driving forces for human Cu transport - from the cell membrane to the Golgi - our strategy involves a range of complementary biophysical experiments on purified proteins, domain constructs and engineered variants. From our studies, we have discovered that (a) the cytoplasmic C-terminus of Ctr1 binds Cu through its HCH motif with a moderate affinity that allows for Cu delivery to Atox1, (b) Atox1 can interact with CCS and exchange Cu implying cross-reactivity between cytoplasmic chaperones, (c) transfer of Cu from Atox1 to metal-binding domains in ATP7B proceeds through Cu-bridged hetero-protein dimers displaying enthalpy-entropy compensation, (d) conformational changes and domain-domain interactions within ATP7B depend on Cu loading status and minute changes in solvent conditions, and (e), in addition to its cytoplasmic chaperone activity, Atox1 may have functionality in the nucleus as it interacted with several DNA-binding proteins in a yeast two-hybrid screen.

Biophysics

Author

Pernilla Wittung Stafshede

Chalmers, Biology and Biological Engineering, Chemical Biology

Biophysical Journal

0006-3495 (ISSN) 1542-0086 (eISSN)

Vol. 110 3 179A-179A

Subject Categories

Biochemistry and Molecular Biology

Biophysics

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Created

10/7/2017