Identification and initial characterisation of a Plasmodium falciparum Cox17 copper metallochaperone.

Copper is an essential micronutrient for all living organisms as an important catalytic co-factor for key enzymes. In higher eukaryotes intracellular copper is distributed by copper metallochaperones. Copper chelators such as neocuproine and tetrathiomolybdate inhibit Plasmodium falciparum erythrocytic development, indicating a requirement for copper by the parasite. A screen of the P. falciparum genome database identified eight potential copper-requiring protein orthologs, including four candidate copper metallochaperones implicated in the delivery of copper to cytochrome-c oxidase. A P. falciparum Cox17 ortholog (PfCox17) was recombinantly expressed and the purified protein bound reduced copper in vitro. PfCox17 was localised to the parasite cytoplasm. Characterisation of plasmodial proteins involved in copper metabolism will help us understand the role of this essential microelement in plasmodial homeostasis.

A Plasmodium falciparum copper-binding membrane protein with copper transport motifs.

BACKGROUND: Copper is an essential catalytic co-factor for metabolically important cellular enzymes, such as cytochrome-c oxidase. Eukaryotic cells acquire copper through a copper transport protein and distribute intracellular copper using molecular chaperones. The copper chelator, neocuproine, inhibits Plasmodium falciparum ring-to-trophozoite transition in vitro, indicating a copper requirement for malaria parasite development. How the malaria parasite acquires or secretes copper still remains to be fully elucidated. METHODS: PlasmoDB was searched for sequences corresponding to candidate P. falciparum copper-requiring proteins. The amino terminal domain of a putative P. falciparum copper transport protein was cloned and expressed as a maltose binding fusion protein. The copper binding ability of this protein was examined. Copper transport protein-specific anti-peptide antibodies were generated in chickens and used to establish native protein localization in P. falciparum parasites by immunofluorescence microscopy. RESULTS: Six P. falciparum copper-requiring protein orthologs and a candidate P. falciparum copper transport protein (PF14_0369), containing characteristic copper transport protein features, were identified in PlasmoDB. The recombinant amino terminal domain of the transport protein bound reduced copper in vitro and within Escherichia coli cells during recombinant expression. Immunolocalization studies tracked the copper binding protein translocating from the erythrocyte plasma membrane in early ring stage to a parasite membrane as the parasites developed to schizonts. The protein appears to be a PEXEL-negative membrane protein. CONCLUSION: Plasmodium falciparum parasites express a native protein with copper transporter characteristics that binds copper in vitro. Localization of the protein to the erythrocyte and parasite plasma membranes could provide a mechanism for the delivery of novel anti-malarial compounds.