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Metabolism of iron in amoebas
Glavanakovová, Marie ; Šuťák, Róbert (advisor) ; Pyrih, Jan (referee)
Ferritins are complex protein structures for iron storage and detoxification. They are typically found in the cytoplasm of the cell; however, they were also identified in mitochondria and chloroplasts. They are distributed among all living organisms but uncommon in non-photosynthetic protists. Surprisingly, a ferritin gene was found in the genome of free-living amoeba Naegleria gruberi. Bioinformatic analysis confirms that it belongs to a group of eukaryotic ferritins. Upon creating antibodies in a laboratory rat, we localized the protein in mitochondria of N. gruberi. The native recombinant protein without the mitochondrial presequence was used for further in vitro experiments. Using growth experiments, we established that the expression of ferritin is dependent on a concentration of iron in cultivation media; however, we were not able to directly prove its ability to bind iron in vitro. We concluded that the function of the ferritin of N. gruberi is most likely related to metabolism of iron. Side experiments were focused on a different protein from the ferritin family - Dps protein in Acanthamoeba castellanii. Despite of a successful creation of antibodies, we were not able to detect the expression of protein neither in total cell lysates nor in subcellular fractions of acanthamoebas under...
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Iron and copper metabolism in marine microalgae of the genus Ostreococcus
Pilátová, Jana ; Šuťák, Róbert (advisor) ; Falteisek, Lukáš (referee)
The smallest free living eukaryote known as Ostreococcus tauri became along with some related species great experimental models for iron uptake research in marine picoplankton. The ecological context of such topic is very interesting considering the nature of adaptation to iron limitation and its connection to the copper metabolism. Our experiments has simulated iron and copper limiting conditions of the ocean, as a control we used iron and copper repleted cultivation media. The maximum cell counts were two- to threefold higher in iron-repleted medium compared to iron- depleted one. There was the only exception showing no difference in growth - O. lucimarinus coming from open ocean with high irradiance levels and very low iron concentrations, which all together made it the best adapted species. The reinoculation of cell cultures after a week cultivation into to the same iron/copper containing media led to unmasking the copper effect on growth, which was much weaker than encountered with iron (again except of O. lucimarinus). Iron sufficiency positively affects heme b and chlorophyll a and b content with no significant copper dependency. The circadian timing of heme b and chlorophyll a and b content shows the increasing trend during the day followed by decrease at night. This might be caused by...
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Function and structure of hydrogenases and their occurence in organisms
Košťálová, Alena ; Hrdý, Ivan (advisor) ; Šuťák, Róbert (referee)
Hydrogenases are microbial enzymes catalyzing reversible oxidation of molecular hydrogen. These enzymes are mostly found in prokaryotes, but a few of them are present in eukaryotes as well. Hydrogenases are metalloproteins which are classified into three classes, [NiFe]-, [FeFe]- and [Fe]-hydrogenases, based on the composition of the active site. [NiFe]-hydrogenases are heterodimeric proteins with the active site localized in the large subunit. [FeFe]-hydrogenases are mostly monomeric and their active site is called H-cluster. Hydrogenases often possess additional domains that contain redox centers, mostly iron-sulfur clusters. Most of the accessory domains of hydrogenases are homologous to other redox complexes, e. g. Complex I of respiratory chain. Maturation of hydrogenases is a complex process involving the activity of several proteins, some of which have been already partially characterized. Compelling evidence indicates that [NiFe]- and [FeFe]-hydrogenases are phylogenetically distinct classes of proteins. This work deals with the differences among hydrogenases including their function, structure, maturation and distribution, and also mentions their similarities with other enzymes.
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Iron as a factor of virulence of parasitic protists
Glavanakovová, Marie ; Šuťák, Róbert (advisor) ; Smutná, Tamara (referee)
Iron is an essential nutrient for metabolism in almost all living organisms. Its importance for many crucial cellular processes originates primarily from the flexibility of available redox potentials. However, the high toxicity of free iron ions is well known. For most organisms, iron is simultaneously and paradoxically essential and toxic. Iron acquisition is crucial for parasitic organisms because it is needed for multiplication in hosts. Many studies have examined the relationship between iron availability and parasite development as a primary factor of virulence. These experiments commonly use chelators, chemical compounds that bind specifically and with high affinity iron ions, and are especially used for iron overload treatment. This thesis summarizes the influence of iron overload or deprivation in the host on the development of parasitic organisms and the impact of chelating agents on the virulence of selected parasitic protists, including the Plasmodium, Leishmania, Trypanosoma, Trichomonas and Tritrichomonas genuses.
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The metabolism of amino acids in parasitic and anaerobic protists
Jakubec, Dávid ; Šuťák, Róbert (advisor) ; Hrdý, Ivan (referee)
Parasitic protists are the cause of countless pathological conditions and economic issues in many parts of the world. While being phylogenetically unrelated, they share many similarities in their approach to satisfying the essential needs. Unlike the much studied energy metabolism, amino acids utilisation pathways are rather unexplored areas. This review shows that in many cases, the parasitic life style has not had the same impact on the amino acid metabolism as it did on the energy metabolism of the protists, which is often severely reduced. Novel pathways have been found in many of the organisms in question, for the biosynthesis of amino acids deemed both essential and non-essential in humans. The arginine dihydrolase pathway found in Trichomonas and Giardia represents a complely new way of utilising the said amino acid. The metabolism of sulfur-containing amino acid has been a matter of intensive research for their non-proteogenic roles. Polyamines are organic nitrogenous compounds involved in many vital processes in the cells, including DNA replication and protein translation. The synthesis of polyamines and their derivatives is elucidated, as it is directly connected to the amino acid metabolism. Finally, the exploitation of the unique pathways described integrates the previous research with the aim...
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