HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplementary data Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Laliberté, J. Articles by Labbé, S. Search for Related Content PubMed PubMed Citation Articles by Laliberté, J. Articles by Labbé, S. Agricola Articles by Laliberté, J. Articles by Labbé, S.

Mechanisms of copper loading on the Schizosaccharomyces pombe copper amine oxidase 1 expressed in Saccharomyces cerevisiae

Département de Biochimie, Université de Sherbrooke, 3001 12e Avenue Nord, Sherbrooke, Québec J1H 5N4, Canada

Correspondence
Simon Labbé
Simon.Labbe{at}USherbrooke.ca

Copper amine oxidases (CAOs) are found in almost every living kingdom. Although Saccharomyces cerevisiae is one of the few yeast species that lacks an endogenous CAO, heterologous gene expression of CAOs from other organisms produces a functional enzyme. To begin to characterize their function and mechanisms of copper acquisition, two putative cao⁺ genes from Schizosaccharomyces pombe were expressed in S. cerevisiae. Expression of spao1⁺ resulted in the production of an active enzyme capable of catalysing the oxidative deamination of primary amines. On the other hand, expression of spao2⁺ failed to produce an active CAO. Using a functional spao1⁺–GFP fusion allele, the SPAO1 protein was localized in the cytosol. Under copper-limiting conditions, yeast cells harbouring deletions of the MAC1, CTR1 and CTR3 genes were defective in amine oxidase activity. Likewise, atx1 null cells exhibited no CAO activity, while ccc2 mutant cells exhibited decreased levels of amine oxidase activity, and mutations in cox17 and ccs1 did not cause any defects in this activity. Copper-deprived S. cerevisiae cells expressing spao1⁺ required a functional atx1⁺ gene for growth on minimal medium containing ethylamine as the sole nitrogen source. Under these conditions, the inability of the atx1 cells to utilize ethylamine correlated with the lack of SPAO1 activity, in spite of the efficient expression of the protein. Cells carrying a disrupted ccc2 allele exhibited only weak growth on ethylamine medium containing a copper chelator. The results of these studies reveal that expression of the heterologous spao1⁺ gene in S. cerevisiae is required for its growth in medium containing ethylamine as the sole nitrogen source, and that expression of an active Schiz. pombe SPAO1 protein in S. cerevisiae depends on the acquisition of copper through the high-affinity copper transporters Ctr1 and Ctr3, and the copper chaperone Atx1.