AbstractMalaria is a serious parasitic disease, caused by unicellular protozoa of the genus Plasmodium, which is responsible for high morbidity and mortality rates among those infected. Only in 2013, 198 million clinical cases of malaria infection and more than 0.5 million deaths were reported.
Although currently confined to tropical and subtropical Countries, the change in climate suitability for transmission of malaria as a result of global warming is associated with a predicted net increase in the global population at risk. In fact, the progress in reducing malaria incidence has been slower than expected in the last decade. Indeed, the fight against malaria is hindered by antimalarial drug resistances arising in the parasite and by the development of mosquitoes that are resistant to pesticides.
Despite more than 20 candidate vaccine constructs are undergoing preclinical development and first trials, a vaccination program is still not available. The scientific community is therefore committed to overcome these issues, developing different antimalarial strategies. The efforts made so far identified the iron homeostasis and the transport mechanisms involved in its regulation process, as a weak point of the intraerythrocytic Plasmodium parasite. In fact, many antimalarials already target the parasite’s iron detoxification pathways. However, there is critical need for a more intimate understanding of aspects of the biochemistry of the malaria parasite to inform future strategies of control. One such aspect that requires more investigation is the physiological role, function, and mechanism of Plasmodium membrane transporter proteins.
Bioinformatic studies of the most virulent human malaria parasite, Plasmodium falciparum, genome identified a putative homologue of the Vacuolar Iron Transporter (VIT) family, subsequently named PfVIT. This transporter was recently observed to conduct ferrous iron transport and it is likely to play a key role for the development of malarial parasites in their host cells. Acquiring a better knowledge on PfVIT function, structure and location is of great interest since it would give precious insights into the iron homeostasis regulation mechanisms of Plasmodium falciparum. Moreover, biophysical information on PfVIT would be fundamental for the study of other transporters belonging to the poorly characterised VIT family.
In this thesis, I present the information regarding PfVIT gained performing various biochemical, molecular, and computational investigations. I have accomplished the heterologous overproduction of PfVIT in Escherichia coli and its purification in functional form. This achievement allowed the functional characterisation of the transporter and the production of antibodies for its subcellular localisation. Moreover, PfVIT structure was modelled using a De-novo method. The analysis of the idealistic PfVIT structure permitted the identification of some residues, possibly constituting the iron binding site.
|Date of Award||Jul 2018|
|Supervisor||Irina Tikhonova (Supervisor), Christopher Law (Supervisor) & John Dalton (Supervisor)|