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Vesicular roles of Arp2/3 nucleation-promoting factors
Dostál, Vojtěch ; Libusová, Lenka (advisor) ; Malínský, Jan (referee) ; Befekadu, Asfaw (referee)
F-actin is involved in key aspects of vesicular traffic, such as membrane deformation, tubulation and vesicle motion. Branching of F-actin is mediated by Arp2/3 but this complex must first be activated by so-called nucleation-promoting factors (NPFs). These factors play an essential role in the decision where and when branched actin should form on the membrane surface. The thesis focuses on the mechanisms which underlie localization and activation of NPFs, especially in terms of the phosphoinositide composition of the vesicle membranes. I show that one of the NPFs, the WASH complex, does not exclusively depend on the retromer complex for its membrane anchoring, as previously theorized. Rather, its understudied subunit SWIP enables the complex to independently bind to the membrane. I also present data showing that the WASH complex has essential roles in maintaining lysosomal function. Additionally, I elucidate the function of another NPF known as WHAMM in the ERGIC compartment, showing that it depends on the presence of myotubularin 9 for its ability to form membrane tubules. The thesis improves our understanding of the interface between the actin cytoskeleton and intracellular membrane system.
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HPLC determination of thiamine in barley, malt and food supplements
Vítková, Kateřina ; Běláková, Sylva (referee) ; Vespalcová, Milena (advisor)
The aim of this diploma thesis is focused on determination of B vitamin in food especially in malting barley and malt. This diploma thesis is consists of theoretical and experimental parts including the conclusion and supplements. Theoretical part describes thiamine characteristic, chemical structure, chemical and physical properties, function of thiamin in organism, impact it´s insufficiency to the human health, occurrence in common food and methods for it´s determination. Further are describing theoretical principles of high-efficiency liquid chromatography, chromatography on reverse phase, derivatization and ways of validation. In the Experimental section is detailed description of used method for determination of thiamine in barley and malt, fresh mash and beer but also in common food supplements as are B-kompex, Pangamin and Thiamin. Determination was made by high-efficiency liquid chromatography on reverse phase with isocratic elution and fluorescent detection. Next part introduce the results of theses, discussion of experimental work, volume of thiamine in analyzed food and validation parameters of used method. In the conclusion are valorized results of experimental work. In the attachment are graphs of standard curves, chromatogram and calculations of validation parameters. Diploma thesis was realized in laboratory of Chemistry and Technology of Foodstuff, Faculty of chemistry, Brno University of Technology.
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