Structure and Cu(I)-binding properties of the N-terminal soluble domains of Bacillus subtilis CopA

Singleton, Chloe, Banci, Lucia, Ciofi-Baffoni, Simone, Tenori, Leonardo, Kihlken, Margaret A., Boetzel, Ruth and Le Brun, Nick E. (2008) Structure and Cu(I)-binding properties of the N-terminal soluble domains of Bacillus subtilis CopA. Biochemical Journal, 411 (3). p. 571. ISSN 0264-6021

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Abstract

CopA, a P-type ATPase from Bacillus subtilis, plays a major role in the resistance of the cell to copper by effecting the export of the metal across the cytoplasmic membrane. The N-terminus of the protein features two soluble domains (a and b), that each contain a Cu(I)-binding motif, MTCAAC. We have generated a stable form of the wild-type two-domain protein, CopAab, and determined its solution structure. This was found to be similar to that reported previously for a higher stability S46V variant, with minor differences mostly confined to the Ser46-containing ß3-strand of domain a. Chemical-shift analysis demonstrated that the two Cu(I)-binding motifs, located at different ends of the protein molecule, are both able to participate in Cu(I) binding and that Cu(I) is in rapid exchange between protein molecules. Surprisingly, UV–visible and fluorescence spectroscopy indicate very different modes of Cu(I) binding below and above a level of 1 Cu(I) per protein, consistent with a major structural change occurring above 1 Cu(I) per CopAab. Analytical equilibrium centrifugation and gel filtration results show that this is a result of Cu(I)-mediated dimerization of the protein. The resulting species is highly luminescent, indicating the presence of a solvent-shielded Cu(I) cluster.

Item Type: Article
Faculty \ School: Faculty of Science > School of Chemistry
Related URLs:
Depositing User: Rachel Smith
Date Deposited: 07 Apr 2011 14:09
Last Modified: 17 Sep 2020 23:38
URI: https://ueaeprints.uea.ac.uk/id/eprint/28489
DOI: 10.1042/BJ20071620

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