Document Type Master's Dissertation Author Nkomo, Ntombikayise Precious email@example.com URN etd-07192011-154844 Document Title iTRAQ-mediated proteome analysis of the response of Plasmodium falciparum to a novel antimalarial compound Degree MSc Department Microbiology and Plant Pathology Supervisor
Advisor Name Title Dr D Mancama Co-Supervisor Prof L-M Birkholtz Co-Supervisor Prof J Theron Supervisor Keywords
- Plasmodium falciparum
Date 2011-04-07 Availability restricted AbstractThe malaria parasite Plasmodium falciparum is one of four species of Plasmodium in the phylum Apicomplexa that infect humans, and is responsible for virtually all deaths from this disease, as well as high levels of morbidity. Malaria has a profound impact on the public health and education systems, economies and social structures of many developing countries, particularly Africa (Sims et al., 2006). Proteomic analysis of this deadly parasite began nearly two decades ago with investigations of 2D gel patterns of parasite proteins before and after the application of antimalarial drugs (Pinswasdi et al., 1987). Current efforts focus on research into novel compounds and on measures to prevent or delay resistance once new drugs are introduced. To develop new antimalarial drugs, ethnobotanical investigation in traditional medicine can be an important source of new leads. African traditional medicine uses numerous plants that can be a source of new antimalarials (Karou et al., 2003). The aim of this research was to develop and employ new protocols in proteomics to investigate the effects on P. falciparum of a novel naturally derived antimalarial compound, Reference #31 (Ref#31). This knowledge contributes towards better understanding the mechanisms underlying the efficacy of Ref#31, and may ultimately be useful to improving the compound‟s further development as an antimalarial. Chapter 1: this chapter introduces the novel antimalarial compound Ref#31, iTRAQ technology, and different techniques that have been used over the years to study the complex proteome of P. falciparum. Chapter 2: describes the different approaches used to optimise iTRAQ technology and maximise the amount of Plasmodium hits identified. Cation exchange and SDS-PAGE were used to fractionate protein samples. Data analysis was performed with Protein Pilot 2.0 (Applied Biosystems). It also describes an in-house approach to improve parasite proteome characterisation, by depleting levels of haemoglobin present in cell culture protein lysates. The approach uses Ni-NTA (Nickel-Nitrilotriacetic acid) agarose (Qiagen), a tetradentate chelating adsorbent. Chapter 3: investigates the morphological effects of the novel plant-derived antimalarial lead Ref#31, on the parasite‟s intraerythrocytic development cycle. Cultured P. falciparum parasites were treated for 48 hrs with different drug concentrations and the parasitemia was determined by Giemsa stain and light microscopy. Blood smears were taken every 6 hr for a period of 48 hrs. The second half of this work describes the analysis of parasite proteins following parasite treatment with Ref#31. Three time points were analysed, 6, 12 and 18 hrs.
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Please cite as follows:
Nkomo, NP 2010, iTRAQ-mediated proteome analysis of the response of Plasmodium falciparum to a novel antimalarial compound, MSc dissertation, University of Pretoria, Pretoria, viewed yymmdd < http://upetd.up.ac.za/thesis/available/etd-07192011-154844/ >
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