Dynamic Multibody Protein Interactions Suggest Versatile Pathways for Copper Trafficking

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• 作者：Aaron M. Keller ; Jaime J. Ben铆tez ; Derek Klarin ; Linghao Zhong ; Matthew Goldfogel ; Feng Yang ; Tai-Yen Chen ; Peng Chen
• 刊名：The Journal of the American Chemical Society
• 出版年：2012
• 出版时间：May 30, 2012
• 年：2012
• 卷：134
• 期：21
• 页码：8934-8943
• 全文大小：536K
• 年卷期：v.134,no.21(May 30, 2012)
• ISSN：1520-5126

文摘

As part of intracellular copper trafficking pathways, the human copper chaperone Hah1 delivers Cu⁺ to the Wilson鈥檚 Disease Protein (WDP) via weak and dynamic protein鈥損rotein 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鈥揗BD3, Hah1鈥揗BD4, and MBD3鈥揗BD4 interaction dynamics within the multidomain system. We found that the two interconverting interaction geometries were conserved in both intermolecular Hah1鈥揗BD and intramolecular MBD鈥揗BD interactions. The Hah1鈥揗BD 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鈥揗BD 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⁺.