title: Dynamic multibody protein interactions suggest versatile pathways for copper trafficking.
abstract: As part of intracellular copper trafficking pathways, the human copper chaperone Hah1 delivers Cu(+) to the Wilson's Disease Protein (WDP) via weak and dynamic protein-protein interactions. WDP contains six homologous metal binding domains (MBDs) connected by flexible linkers, and these MBDs all can receive Cu(+) from Hah1. The functional roles of the MBD multiplicity in Cu(+) trafficking are not well understood. Building on our previous study of the dynamic interactions between Hah1 and the isolated fourth MBD of WDP, here we study how Hah1 interacts with MBD34, a double-domain WDP construct, using single-molecule fluorescence resonance energy transfer (smFRET) combined with vesicle trapping. By alternating the positions of the smFRET donor and acceptor, we systematically probed Hah1-MBD3, Hah1-MBD4, and MBD3-MBD4 interaction dynamics within the multidomain system. We found that the two interconverting interaction geometries were conserved in both intermolecular Hah1-MBD and intramolecular MBD-MBD interactions. The Hah1-MBD interactions within MBD34 are stabilized by an order of magnitude relative to the isolated single-MBDs, and thermodynamic and kinetic evidence suggest that Hah1 can interact with both MBDs simultaneously. The enhanced interaction stability of Hah1 with the multi-MBD system, the dynamic intramolecular MBD-MBD interactions, and the ability of Hah1 to interact with multiple MBDs simultaneously suggest an efficient and versatile mechanism for the Hah1-to-WDP pathway to transport Cu(+).
authors.affiliation: Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, USA.
authors.forename: Aaron M
authors.initials: AM
authors.lastname: Keller
authors.forename: Jaime J
authors.initials: JJ
authors.lastname: Benítez
authors.forename: Derek
authors.initials: D
authors.lastname: Klarin
authors.forename: Linghao
authors.initials: L
authors.lastname: Zhong
authors.forename: Matthew
authors.initials: M
authors.lastname: Goldfogel
authors.forename: Feng
authors.initials: F
authors.lastname: Yang
authors.forename: Tai-Yen
authors.initials: TY
authors.lastname: Chen
authors.forename: Peng
authors.initials: P
authors.lastname: Chen
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D001692:Biological Transport; D027682:Cation Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D003300:Copper / Q000378:metabolism*; D000073840:Copper-Transporting ATPases; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007700:Kinetics; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary
chemical_list: D027682:Cation Transport Proteins; D003300:Copper; D000251:Adenosine Triphosphatases; D000073840:Copper-Transporting ATPases