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Effect of selected compounds on NMPylation of nsp9 SARS-CoV-2 catalyzed by NiRAN domain of nsp12 SARS-CoV-2
Fedák, Michal ; Šilhán, Jan (advisor) ; Vaněk, Ondřej (referee)
SARS-CoV-2 pandemic that begun in the end of 2019 caused over 770 milion confirmed cases of infection and almost 7 milion deaths worliwide. Despite pandemic being suppressed thanks to vaccines and restriction measures, the research of new medication targeting this virus continues. One of the possible targets of SARS-CoV-2 is nonstructural protein 9 (nsp9), which is essential for virus repliation. Just recently it was revealed that this protein can covalently bind nukleoside monophosphate (NMPylation) or RNA chain of various length (RNAylation). These reactions are catalyzed by nidovirus RNA-dependent RNA polymerase (RdRp) asociated nucleotidyltransferase (NiRAN) domain of nonstuctural protein 12 (nsp12). RNAylation of nsp9 showed to be crucial step in synthesis of RNA cap. The function of NMPylation was not jet revealed, but it is believed that this modification of nsp9 could play a role in priming of RNA synthesis. Some studies have already investigated inhibition of NMPylation. In this diploma thesis recombinant nsp9 was prepared and used in NMPylation reactions. Negative effect of DMSO on nsp9 NMPylation was observed. Three compounds (saquinavir, darunavir and conivaptan) selected based on the results of in silico studies did not show ability to inhibit NMPylation of nsp9. Next, it was confirmed...
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Termín a způsob aplikace klasické a inhibované močoviny ve výživě kukuřice
Miloš, Lukáš
The aim of this bachelor thesis was to examine the effect of mineral fertilizers application on the grain yield of corn. The 4-year field experiment was conducted in years 2019-2022 in the form of small plot field experiment at the Field experimental station Žabčice. The primary objective of the experiment was to compare the method and term of application, as experimental fertilizers were chosen classic urea and its fully inhibited variant, thus urea with nitrification (IN) and urease (IU) inhibitor. The following treatments were included in the experiment: 1. Urea + IN + IU to the depot (autumn), 2. Urea to the depot (autumn), 3. Urea + IN + IU to the depot (spring), 4. Urea to the depot (spring), 5. Urea + IN + IU wide (spring), 6. Urea wide (spring).The work also focuses on average comparison of term (autumn, spring) and method of application (to the depot, wide), regardless of investigated fertilizer as well as to the average yield of grain after the application of the examined fertilizers (urea, urea + IN + IU). The average four-year results showed a difference of 4.3 % in grain yield after the application of urea fertilizer (8.38 t/ha), regardless of the term and method of application, compared to its inhibited variant (8.03 t/ha). The application of the examined fertilizers in autumn resulted in an increase in average grain yield by 5.8% compared to application in spring. The last factor investigated was the method of application, with maize fertilised widely yielding 10.3% higher compared to depot fertilisation.
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Design and evaluation of potential viral methyltransferase inhibitors
Kocek, Hugo ; Nencka, Radim (advisor) ; Grantz Šašková, Klára (referee)
A global pandemic of SARS-CoV-2 confirmed the pandemic potential of the Coronaviridae family and pointed out the need for novel antiviral drugs. The SARS-CoV-2 pandemic has been tamed thanks to mRNA vaccines; however, monoclonal antibodies and small molecules such as nirmatrelvir (protease inhibitor), remdesivir (polymerase inhibitor), or molnupiravir (mutagen) are currently also available. It is worth noting that remdesivir and molnupiravir were previously investigated as antivirals against different pathogens. SARS-CoV-2 encodes 16 non-structural proteins, and two of them - methyltransferases (MTases) nsp14 and nsp16 - participate in RNA capping as the virus must mimic the host's mRNA to evade the cellular antiviral sensors (e.g., IFIT1) and replicate. These MTases are structurally very similar to those of SARS-Co-V; therefore, we might expect that inhibitors of SARS-CoV-2 MTases could be used in the future against different coronaviruses. For the reasons mentioned above, this thesis focuses on developing novel MTase inhibitors targeting SARS-CoV-2 nsp14 and nsp16. The design was based on S-adenosyl-L-homocysteine (SAH; endogenous inhibitor of MTases) and an in silico compound library was constructed with various replacements for SAH's amino acid moiety. The potential inhibitory activity was...
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Synthesis of peptidic inhibitors targeting PA-PB1 interface of influenza RNA polymerase
Palacková, Miroslava ; Machara, Aleš (advisor) ; Veselý, Jan (referee)
The submitted Thesis deals with preparation of a hexapeptides inhibiting protein-protein interaction of PA-PB1 subunits of influenza RNA polymerase. Crucial part of the Thesis represent modifications of particular small hexapeptide at its two "hot spots". It means at positions that significantly contribute to the binding of both subunits. These modifications resulted in preparation of two series of distinct hexapeptides. With regards to the fact that one designed hexapeptide contains unnatural and commercially unavailable amino acids this amino acid had to be prepared from simple building blocks. Apart from aforementioned work the Thesis also covers effort to prepared bicyclic peptide that contains sequences of peptidic inhibitor of protein-protein interactions and also cell-penetration peptide. Key words: synthesis, peptides, inhibitors, influenza, polymerase
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The Role of B-cell Receptor Signaling in Lymphoid Malignancies
Kochmannová, Kateřina ; Havránek, Ondřej (advisor) ; Štěpánek, Ondřej (referee)
The aim of this thesis is to review current knowledge about tumor-specific B cell receptor (BCR) signaling and related novel therapy options in B-cell malignancies with the main focus on non-Hodgkin lymphomas (NHL). To a certain degree, the pathogenic BCR signaling mirrors normal forms of BCR signaling, antigen-induced and tonic. Differences between antigen- dependent and antigen-independent forms of BCR signaling are well characterized in two major subtypes of diffuse large B-cell lymphoma, the most common type of NHL. In addition to the conventional chemotherapy, several BCR inhibitors targeting BTK and PI3K have been approved for the treatment of lymphoid malignancies. However, improvements in the tumor specificity, toxicity profiles and patients selection are needed. A better understanding of BCR signaling deregulation and overall tumor pathogenesis is believed to further improve NHL treatment outcomes. Keywords: B-cell malignancies, non-Hodgkin lymphoma, B-cell receptor, tumor signaling, targeted therapy, inhibitors
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Vliv stabilizovaného dusíkatého hnojiva se sírou na obsah minerálního dusíku v půdě a na výnos a kvalitu semen řepky ozimé
Pliska, Radim
The thesis deals with the influence of application of stabilized nitrogen fertilizer with sulphur and other nitrogenous fertilizers, on the content of mineral nitrogen in the soil. The yield and oiliness of winter rape seeds is also evaluated. The problem was solved in the agricultural year 2015/2016 as a small-plot field experiment at the experimental station in Žabčice u Brna (179 m.) and at the research fodder station Vatín near Žďár nad Sázavou (560 m.). The following 5 variants of the fertilizer were included in the experiment: LAD regenerative variant (104 kg N/ha), DASA regenerative variant (104 kg N/ha), ENSIN regenerative variant (136 kg N/ha). In the LAD + DASA and LAD + ENSIN variants, was used for the regenerative fertilizin the LAD fertilizer (51 kg N/ha respectively 64 kg N/ha), DASA (104 kg N/ha) and ENSIN (130 kg N/ha) were applied in the first production fertilizing. Beside the ENSIN variant, it was used for fertilization in I. production fertilizing and II. production fertilizing at a total nitrogen dose of 194 kg N/ha DAM 390 fertilizer. In the ENSIN Different effects of fertilizers on yield were expected, but a statistically significant increase in yields was shown only in the DASA variant, as well as a different effect on oiliness was expected, but there was no statistically significant variant, thought the effect of fertilizer with sulfur, appeared where by all sulfur variant werw increased oilyness compared to the LAD variant. The mineral nitrogen evaluation showed the fertilizer effect with the ENSIN inhibitor, where the ammonium nitrogen ratio exceeded over the nitrate form.
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Vliv přídavku inhibitorů do hnojiva DAM 390 na výnos a olejnatost semen řepky ozimé
Špaňhelová, Klára
Subject of diploma thesis is assessment of influence addition inhibitors in liquid urea–ammonium nitrate (UAN) fertilizer on the yield and oiliness of rape. The experiment was solved in two-years (2017, 2018) form small-field experiment in research station Žabčice. In experiment was included four variants of fertilization: 1. ANCS (ammonium nitrate with calcium sulfate) + UAN (urea ammonium nitrate), 2. ANCS +UAN + IN (nitrification inhibitor), 3. ANCS + UAN + IU (urease inhibitor), 4. ANCS + UAN + IN + IU. Addition of inhibitors in liquid fertilizer UAN didn't has statically conclusive effect on rape seed yield. The highest yield was achieved in variant ANCS + UAN with yield of 4,1 t/ha. The lowest yield was achieved in variant ANCS + UAN + IN with yield 3,9 t/ha. Weather was above average warm and dry in both reference years. The efficiency of nitrification inhibitors with rising temperature decreases linearly. The found oiliness of rape seeds from individual fertilization variants was relatively balanced. The applied inhibitors didn't significantly affect the oil content in the seeds. The highest oiliness was achieved in variant ANCS + DAM with oiliness of 40,7 %. The lowest oiliness was achieved in variant ANCS + UAN + IN + IU with oiliness of 40,4 %.
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Indoleamine 2,3-dioxygenase 1 inhibitors in cancer immunotherapy
Muffová, Barbora ; Poláková, Ingrid (advisor) ; Šroller, Vojtěch (referee)
The indoleamine 2,3-dioxygenase (IDO 1) enzyme is expressed in small amounts in most of the mammalian tissues, and its production is detected also in various types of tumours. IDO 1 catalyses the very first step of the kynurenine pathway, the tryptophan conversion to N-formylkynurenine, which is further metabolized to kynurenine (Kyn). The kynurenine/ tryptophan ratio (kyn/trp) may be used as a prognostic marker in research and treatment of IDO 1+ tumours. The kyn/trp demonstrates the activity of IDO 1 in tumours. The goal of cancer immunotherapy based on IDO 1 inhibition is to reverse or reduce the protumour effects of IDO 1, such as avoiding NK and T cells inhibition and activation of regulatory T cells or association with tumour-associated macrophages (TAM). IDO 1 inhibitors have been examined alone in monotherapy or together with cytotoxic T-lymphocytes antigen 4 (CTLA-4) inhibitors and programmed cell death protein 1 (PD-1) inhibitors in combined therapy. Recently, several studies are dedicated to invent inhibitors, which are able to inhibit the activity of other trp-catalysing enzymes, the indoleamine 2,3-dioxygenase 2 (IDO 2) and tryptophan 2,3-dioxygenase (TDO), together with the IDO 1 activity. Cancer immunotherapy based on IDO 1 inhibition may be combined also with chemotherapy.
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Investigation and inhibition of α-synuclein aggregation
Afitska, Kseniia ; Yushchenko, Dmytro (advisor) ; Žídek, Lukáš (referee) ; Bařinka, Cyril (referee)
α-Synuclein (AS) is a small intrinsically disordered protein expressed in neurons and abundantly present in synapses where it is involved in regulation of synaptic vesicle-mediated protein trafficking. Misfolding of AS into amyloid fibrils is a key process in progression of Parkinson's disease (PD), the second most common neurodegenerative disorder which has no cure to date. Inhibition of AS aggregation and blocking of cell-to-cell spreading of AS fibrils is a promising strategy for PD treatment. However, rational design of inhibitors of this type remains complicated due to the lack of thorough knowledge about the mechanisms of aggregation. Therefore, the aim of this thesis was to gain deeper knowledge about AS aggregation and to apply it for developing inhibitors of AS fibrillization. In my work on the mechanisms of AS aggregation, I first determined that the concentration of AS that enables the fibril growth is an order of magnitude lower than the concentration of AS required for initial fibril formation from monomers. I explored fibril disaggregation at AS concentrations below its Kd value, and characterized AS aggregation at low micromolar concentrations. I then investigated how different modifications of AS C-terminus (namely, extensions of various sizes and charges) affect fibril growth and...
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