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Různé metodické přístupy za účelem snížení exprese podjednotek komplexu I u hmyzích forem \kur{Trypanosoma brucei}
HEROUTOVÁ, Barbora
Complex I (NADH:ubiquinone dehydrogenase) is the largest protein complex of the mitochondrial electron transport chain, but its presence and activity are not essential for the growth of two life cycle stages of Trypanosoma brucei. Here, we implemented various genetic methods to generate T. brucei cell lines that will lack or will have decreased levels of this complex. The methodological approaches included i) RNA interference of NDUFA6, a subunit predicted to be essential for the structural integrity, and of NUBM, a subunit essential for the complex I activity; ii) generation of double knock-out of NDUFA6 using CRISPR/Cas9; iii) generation of double knock-out of NUBM using homology replacement. These generated tools will help us to elucidate if this highly conserved complex is essential for other life cycle stages of this medically important parasite
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Complex I of mitochondrial respiratory chain a its disorders.
Rodinová, Marie ; Hansíková, Hana (advisor) ; Kalous, Martin (referee)
NADH: ubiquinone oxidoreductase (Complex I) is a multisubunit protein complex of inner mitochondrial membrane. Complex I is the biggest and most complicated part of oxidative phosphorylation system, which is responsible for the cell ATP production. It consists of 45 subunits. 7 subunits are mitochondrial encoded, remainder 38 are nuclear encoded. NADH: ubiquinone oxidoreductase has L-shaped structure, which is built of two arms: membrane arm and matrix located peripheral arm. Complex I oxidize the NADH molecule. The electron transport is coupled with proton pumping across the inner mitochondrial membrane to intermembrane space, where proton gradient developed and which is used by ATP synthase to ATP synthesis. Deficiencies of NADH: ubiquinone oxidoreductase represent extensive, clinically and genetic heterogeneous group of mitochondrial diseases. Decrease of activity and amount of complex I, decrease of ATP production, changes of membrane potential, mitochondrial morphology and mitochondrial network and increasing of production of reactive oxygen species are found in cells with defects of NADH: ubiquinone oxidoreductase. Combination of this features lead to serious illnesses, which are almost fatal and we still haven't any useful therapy. Aim of this study is to summarize present knowledge about...
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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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Complex I of mitochondrial respiratory chain a its disorders.
Rodinová, Marie ; Kalous, Martin (referee) ; Hansíková, Hana (advisor)
NADH: ubiquinone oxidoreductase (Complex I) is a multisubunit protein complex of inner mitochondrial membrane. Complex I is the biggest and most complicated part of oxidative phosphorylation system, which is responsible for the cell ATP production. It consists of 45 subunits. 7 subunits are mitochondrial encoded, remainder 38 are nuclear encoded. NADH: ubiquinone oxidoreductase has L-shaped structure, which is built of two arms: membrane arm and matrix located peripheral arm. Complex I oxidize the NADH molecule. The electron transport is coupled with proton pumping across the inner mitochondrial membrane to intermembrane space, where proton gradient developed and which is used by ATP synthase to ATP synthesis. Deficiencies of NADH: ubiquinone oxidoreductase represent extensive, clinically and genetic heterogeneous group of mitochondrial diseases. Decrease of activity and amount of complex I, decrease of ATP production, changes of membrane potential, mitochondrial morphology and mitochondrial network and increasing of production of reactive oxygen species are found in cells with defects of NADH: ubiquinone oxidoreductase. Combination of this features lead to serious illnesses, which are almost fatal and we still haven't any useful therapy. Aim of this study is to summarize present knowledge about...
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