La très catastrophique visite du zoo

Joël Dicker

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Doctoral Thesis / Dissertation from the year 2014 in the subject Biology - Diseases, Health, Nutrition, grade: 1.3, University of Marburg (European virtual Institute for Malaria Research), language: English, abstract: This study focuses on the discovery of an alternative secretory pathway to the ER/Golgi route in the malaria parasite P. falciparum in infected RBCs. Two proteins appeared to be promising candidates of an alternative secretory pathway: the PfADP-ribosylation factor 1 (ARF1) and the Pfadenylate kinase 2 (AK2). Both proteins contained an N-myristoylation site at their N-terminus, which is indicative for N-myristoylation. N-myristoylation is a co-translational modification of a protein, whereby a fatty acid (myristate) is irreversibly attached to the glycine residue at the N-terminus of a protein via the PfN-myristoyltransferase (NMT). A preceding proteomic analysis of the parasitophorous vacuole and a reporter construct study proposed for both PfARF1 (determined by a proteomic study) and PfAK2 (determined by a reporter construct study) PV localization although both proteins lacked a signal peptide. That's why it was hypothesized whether or not N-myristoylation would drive protein secretion across the parasite plasma membrane (PPM). The subcellular localization of the PfARF1/GFP parasites and the PfAK2/GFP parasites, respectively, were analyzed via epifluorescence microscopy and biochemical methods. In parallel, another batch of reporter constructs were generated and analyzed, where the N-myristoylation site of PfARF1 (this study) and PfAK2 (Ma et al., 2012), respectively, was removed (PfARF1G2A/GFP and PfAK2G2A/GFP). Live cell imaging showed that the fusion protein ARF1/GFP was localized as dot-like structures in the parasite. In contrast, the phenotype of the fusion protein of the PfARF1G2A/GFP parasites showed an evenly distributed signal in the parasite cytosol. Further analysis of the subcellular localization of the PfARF1 strongly supports its loc

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