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Volume 148, 2011
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Relating dynamic protein interactions of metallochaperones with metal transfer at the single-molecule level

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Abstract

Metallochaperones undertake specific interactions with their target proteins to deliver metal ions inside cells. Understanding how these protein interactions are coupled with the underlying metal transfer process is important, but challenging because they are weak and dynamic. Here we use a nanovesicle trapping scheme to enable single-molecule FRET measurements of the weak, dynamic interactions between the copper chaperone Hah1 and the fourth metal binding domain (MBD4) of WDP. By monitoring the behaviors of single interacting pairs, we visualize their interactions in real time in both the absence and the presence of various equivalents of Cu1+. Regardless of the proteins' metallation state, we observe multiple, interconverting interaction complexes between Hah1 and MBD4. Within our experimental limit, the overall interaction geometries of these complexes appear invariable, but their stabilities are dependent on the proteins' metallation state. In apo–holo Hah1–MBD4 interactions, the complexes are stabilized relative to that observed in the apo–apo interactions. This stabilization is indiscernible when Hah1′s Cu1+-binding is eliminated or when both proteins have Cu1+ loaded. The nature of this Cu1+-induced complex stabilization and of the interaction complexes are discussed. These Cu1+-induced effects on the Hah1–MBD4 interactions provide a step toward understanding how the dynamic protein interactions of copper chaperones are coupled with their metal transfer function.

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Article information


Submitted
07 Apr 2010
Accepted
14 Apr 2010
First published
23 Aug 2010

Faraday Discuss., 2011,148, 71-82
Article type
Paper

Relating dynamic protein interactions of metallochaperones with metal transfer at the single-molecule level

J. J. Benítez, A. M. Keller, D. L. Huffman, L. A. Yatsunyk, A. C. Rosenzweig and P. Chen, Faraday Discuss., 2011, 148, 71
DOI: 10.1039/C004913A

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