Das Selenoproteom des Malaria-Erregers Plasmodium falciparum
Final Report Abstract
Many selenoproteins represent important components of cellular redox networks. Most selenocysteine-containing proteins are redox-active, possess in comparison with their cysteine containing orthologs a higher catalytic activity, and are particularly reactive towards electrophilic inhibitors. In the malaria parasite Plasmodium the genes of four putative selenoproteins (PfSel1-4) as well as the genes for a complete selenoprotein biosynthesis machinery have been identified. Within the frame of our DFG-funded project, the genes of all four selenoproteins of Plasmodium falciparum were cloned and heterologously overexpressed as cysteine mutants in Escherichia coli. As far as possible, the recombinant proteins were purified via affinity and size exclusion chromatography. The proteins were studied biochemically, in functional assays and with respect to their oligomerization behavior. For all four selenoproteins, detailed bioinformatics analyses were carried out and indicated similarities between PfSel1 and SelK, PfSel2 and SelT, and between PfSel4 and SelS, whereas PfSel3 remained unique. Furthermore, GFP fusion constructs were generated and, after transformation of Plasmodium falciparum, the subcellular localization was determined for all four proteins using fluorescence microscopy. The data indicate that PfSel3 seems to be located in the nucleus whereas the other three selenoproteins localize to the endoplasmic reticulum. Functional assays indicate roles of PfSel1 and PfSel4 as redox active proteins. Overexpression of PfSel2 in Plasmodium falciparum leads to an increase of free cytosolic calcium. Profiles of the transcripts of the four selenoproteins and other plasmodial redox active proteins were prepared after supplementing the cell culture with sodium selenite, paraquat, sodium nitroprusside, and methylene blue, respectively. The results indicate that the transcripts of the selenoproteins are regulated by the availability of selenium. Moreover, PfSel1, PfSel3, and PfSel4 are down-regulated under nitrosative and oxidative stress. Furthermore the regulation of PfSel1 transcripts under treatment of malaria parasites with coruleoellagic acid, derived from the redox-active and chemopreventive agent ellagic acid, was studied. Based on a collaboration, novel selenium and tellurium containing multifunctional agents interacting with several components of redox metabolism were tested and found to be active against malaria parasites. In addition, sulfur nanoparticles were developed and assessed in a collaborative approach, and determined to exhibit antiplasmodial activity. Last but not least, first results for knocking out the selenoprotein biosynthesis machinery were obtained. These studies will be followed up in the near future.
Publications
- (2009). Compounds structurally related to ellagic acid show improved antiplasmodial activity. Antimicrob. Agents Chemother. 53: 622-630
Sturm N, Hu Y, Zimmermann H, Fritz-Wolf K, Wittlin S, Rahlfs S, Becker K
- (2009). Exploring synthetic avenues for the effective synthesis of selenium- and tellurium-containing multifunctional redox agents. Org. Biomol. Chem. 7:4753-62
Mecklenburg S, Shaaban S, Ba LA, Burkholz T, Schneider T, Diesel B, Kiemer AK, Röseler A, Becker K, Reichrath J, Stark A, Tilgen W, Abbas M, Wessjohann LA, Sasse F, Jacob C
- (2011). Selective antimicrobial activity associated with sulfur nanoparticles. J. Biomed. Nanotech. 7, 1–11
Schneider T, Baldauf A, Ba LA, Jamier V, Khairan K, Sarakbi MB, Reum N, Schneider M, Röseler A, Becker K, Burkholz T, Winyard PG, Kelkel M, Diederich M, Jacob C
