title: N-terminal positively charged amino acids, but not their exact position, are important for apicoplast transit peptide fidelity in Toxoplasma gondii.
abstract: The non-photosynthetic plastid - or apicoplast - of Toxoplasma gondii and other apicomplexan parasites is an essential organelle and promising drug target. Most apicoplast proteins are encoded in the nucleus and targeted into the organelle through the apicoplast's four membranes courtesy of a bipartite N-terminal leader sequence comprising of an endomembrane signal peptide followed by a plastid transit peptide. Apicoplast transit peptides, like plant plastid transit peptides, have no primary consensus, are variable in length and may be distinguishable only by a relative depletion of negative charged residues and consequent enrichment in basic residues. In this study we examine the role of charged residues within an apicoplast transit peptide in T. gondii by point mutagenesis. We demonstrate that positive charged residues, combined with the absence of negatively charged amino acids, are essential for apicoplast transit peptide fidelity, as also observed in P. falciparum. Furthermore, we show that positive charge is more important at the transit peptide's N-terminus than its C-terminus, and that the nature of the positive residue and the exact position of the N-terminal positive charge are not important. These results suggest that a simple, rule-based prediction for T. gondii transit peptides, similar to that successfully implemented for P. falciparum should help to identify apicoplast proteins and facilitate the identification of drug targets in this important human pathogen.
authors.affiliation: Plant Cell Biology Research Centre, The School of Botany, The University of Melbourne, Parkville, Melbourne, Victoria 3010, Australia.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Tonkin
authors.forename: David S
authors.initials: DS
authors.lastname: Roos
authors.forename: Geoffrey I
authors.initials: GI
authors.lastname: McFadden
mesh_terms: D000213:Acyl Carrier Protein / Q000378:metabolism; D000595:Amino Acid Sequence; D024361:Amino Acids, Basic; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000469:parasitology; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010455:Peptides / Q000378:metabolism*; D018087:Plastids / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000235:genetics; D021381:Protein Transport / Q000502:physiology; D015800:Protozoan Proteins / Q000378:metabolism*; D014122:Toxoplasma / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D000213:Acyl Carrier Protein; D024361:Amino Acids, Basic; D001426:Bacterial Proteins; D008164:Luminescent Proteins; D010455:Peptides; D021382:Protein Sorting Signals; D015800:Protozoan Proteins; C054562:yellow fluorescent protein, Bacteria