Distinct mechanisms for Ctr1-mediated copper and cisplatin transport

Devis Sinani, David J. Adle, Heejeong Kim, Jaekwon Lee

Research output: Contribution to journalArticle

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Abstract

The Ctr1 family of integral membrane proteins is necessary for high affinity copper uptake in eukaryotes. Ctr1 is also involved in cellular accumulation of cisplatin, a platinum-based anticancer drug. Although the physiological role of Ctr1 has been revealed, the mechanism of action of Ctr1 remains to be elucidated. To gain a better understanding of Ctr1-mediated copper and cisplatin transport, we have monitored molecular dynamics and transport activities of yeast Saccharomyces cerevisiae Ctr1 and its mutant alleles. Co-expression of functional Ctr1 monomers fused with either cyan or yellow fluorescent protein resulted in fluorescence resonance energy transfer (FRET), which is consistent with multimer assembly of Ctr1. Copper near the K m value of Ctr1 enhanced FRET in a manner that correlated with cellular copper transport. In vitro cross-linking of Ctr1 confirmed that copper-induced FRET reflects conformational changes within pre-existing Ctr1 complexes. FRET assays in membrane-disrupted cells and protein extracts showed that intact cell structure is necessary for Ctr1 activity. Despite Ctr1-dependent cellular accumulation, cisplatin did not change Ctr1 FRET nor did it attenuate copper-induced FRET. A Ctr1 allele defective in copper transport enhanced cellular cisplatin accumulation. N-terminal methionine-rich motifs that are dispensable for copper transport play a critical role for cisplatin uptake. Taken together, our data reveal functional roles for structural remodeling of the Ctr1 multimeric complex in copper transport and suggest distinct mechanisms employed by Ctr1 for copper and cisplatin transport.

Original languageEnglish (US)
Pages (from-to)26775-26785
Number of pages11
JournalJournal of Biological Chemistry
Volume282
Issue number37
DOIs
StatePublished - Sep 14 2007

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Cisplatin
Copper
Fluorescence Resonance Energy Transfer
Yeast
Alleles
Molecular Dynamics Simulation
Cell membranes
Platinum
Cell Extracts
Eukaryota
Methionine
Saccharomyces cerevisiae
Molecular dynamics
Assays
Membrane Proteins
Proteins
Yeasts
Monomers
Membranes
Pharmaceutical Preparations

ASJC Scopus subject areas

  • Biochemistry

Cite this

Distinct mechanisms for Ctr1-mediated copper and cisplatin transport. / Sinani, Devis; Adle, David J.; Kim, Heejeong; Lee, Jaekwon.

In: Journal of Biological Chemistry, Vol. 282, No. 37, 14.09.2007, p. 26775-26785.

Research output: Contribution to journalArticle

Sinani, Devis ; Adle, David J. ; Kim, Heejeong ; Lee, Jaekwon. / Distinct mechanisms for Ctr1-mediated copper and cisplatin transport. In: Journal of Biological Chemistry. 2007 ; Vol. 282, No. 37. pp. 26775-26785.
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