The focus of our lab is to further understand the mitochondrial metabolism in trypanosomatid protozoa. T. brucei and Leishmania major are used in the lab as model organisms to elucidate the unique characteristics of each parasitic respiratory chain. Through the use of genetic and biochemical methodology, we hope to identify new drug targets that will create exciting opportunities for drug development to replace the few existing toxic drugs available for these devastating diseases.
Acestor N., Zíková A., Dalley R.A., Anupama A., Panigrahi A.K., Stuart K.D. (2011) Trypanosoma brucei Mitochondrial Respiratome: Composition and Organization in Procyclic Form. Molecular & Cellular Proteomics 10: 9.
F0F1-ATP synthase plays an essential role in Trypanosoma brucei, an important pathogen of humans and livestock that alternates between an insect vector and a mammalian host. Interestingly, the function of the mitochondrial (mt) F0F1-ATP synthase differs between the insect and mammalian stages, producing ATP in the former while consuming ATP to maintain the mt membrane potential in the latter. Purification of this complex revealed 18 subunits, of which 12 possess no obvious homology to any other proteins. The proposed aim of this project is to resolve the function of these novel subunits within the F0F1-ATP synthase. We will determine if these proteins are involved in the structural organization of the complex, in its oligomerization and/or in the regulation of the different enzymatic activities found during the two life cycle stages. We will also elucidate the specific interaction between these proteins within the F0 or F1 moieties. The obtained results will expand our knowledge about this intriguing complex and may open new opportunities for drug development.
The FoF1-ATP synthase can work in reverse, basically hydrolyzing ATP to pump protons across the inner mt membrane. Such an activity is well documented in anaerobic bacteria, but demonstrated cases are rare in eukaryotes and usually involve hypoxic or anoxic conditions. For instance, during ischemic conditions the mammalian FoF1-ATP synthase switches to its hydrolytic state to maintain the essential mt membrane potential, but in doing so it depletes the tissue of ATP and leads to cell death. This ATPase activity is regulated by the inhibitor factor IF1, a small mitochondrial protein. We are investigating if the Trypanosoma homolog of the mammalian IF1, TbIF1, is also involved in the regulation of the ATP synthase activities in T. brucei cells. Further, we need to understand the precise mode of binding and the mechanism of inhibition between TbIF1 and F1-ATPase, which may lead to identification of the interaction interface that can be targeted by structure-based drug design in future.
MIX is a unique mitochondrial (mt) protein found only within the order Kinetoplastida, which includes the medically important parasites Trypanosoma spp. and Leishmania spp. Importantly, MIX is essential for cell viability and even the deletion of one MIX allele leads to the loss of L. major virulence. Furthermore, studies from T. brucei revealed that MIX is associated with several subunits of cytochrome c oxidase, implying a role in the mt respiration of Kinetoplastida. Our main goal is to resolve the primary function of the MIX protein within the cytochrome c oxidase of T. brucei and L. major. The obtained results will expand our knowledge of the unique mitochondrial respiratory machinery in these parasitic protozoa and may illuminate the cyt c oxidase complex as a drug target to treat leishmaniasis.
Moreover, in collaboration with Silvie Trantírková, Phd. we are interested in putative mitochondrial methyltransferase. Methyltranferases are ubiquitous enzymes essential for many cell processes. The main aim of this project is to perform functional analysis of the putative RNA methyltransferase MT420 via RNA interference or double knock-out strategies and to identify its substrates. These studies will illuminate importance of this protein for the viability of the parasite and elucidate its function in methylation processes e.g.mitochondrial ribosomal RNAs, mitochondrial mRNAs or tRNAs. This project will contribute to our understanding about the importance of mitochondrial RNA methylation.
A large collection of helminths is available for comparative studies...
