Blocking helix formation without blocking organellar localization in Plasmodium falciparum

Abstract

Transit peptide (TP) recognition in mitochondria and chloroplast localization is well described and requires a receptor to recognize the TP bound as an amphipathic alpha helix. This functional interaction leads to organellar import of the payload protein. Plasmodium falciparum (Pf), the causative agent of malaria, contains an organelle called the apicoplast. The apicoplast is evolutionarily related to the chloroplast, is essential to the metabolism of Pf, and contains numerous putative drug targets. As in chloroplasts, nuclear-encoded apicoplast proteins must be post-translationally targeted to the apicoplast. In contrast to chloroplast localization, molecular details of TP recognition in Pf are currently unknown. To assess if apicoplast TPs must form helical intermediates for proper organellar localization, we have examined the TP of Pf acyl carrier protein by circular dichroism (CD), nuclear magnetic resonance (NMR), and epifluorescent microscopy of mutant TP-GFP fusions. CD and NMR of acyl carrier protein with its TP in solution are consistent with the presence of a small population of helix in the TP. However, structure-disrupting proline mutations are correctly targeted to the apicoplast when observed in vivo. This observation contradicts the theory that apicoplast TP recognition occurs via a mechanism similar to chloroplast TPs, and instead suggests that the dominant population of disordered TP may be the active form and that Pf has evolved a distinct solution to the problem of organellar targeting.