National Repository of Grey Literature 6 records found  Search took 0.01 seconds. 
Study of intermolecular interactions by isothermal titration calorimetry
Pšenáková, Katarína ; Obšil, Tomáš (advisor) ; Alblová, Miroslava (referee)
Calorimetric methods are used to studying the mechanism of regulation and control of biological processes at the molecular level. Isothermal titration calorimetry (ITC) is used for monitoring the heat exchange of molecular interactions in an environment with constant temperature. This method is also preferred because of its ability of direct measurement of thermodynamic parameters associated with complex formation. ITC measures mostly the energetics of biochemical reactions or molecular interactions protein-protein, protein-ligand, or enzyme-substrate. Main goals of this bachelor thesis were: (I) to acquire working skills with nano isothermal titration calorimeter; (II) to prepare the DNA-binding domain of human transcription factor FOXO4 (FOXO4-DBD); and (III) to study thermodynamic aspects of the interaction between FOXO4-DBD and the double-stranded DNA containing specific binding motif. FOXO4 protein is one of the four members of a "O" subgroup of forkhead transcription factors FOX, which play an important role in many cellular processes including stress resistance, age control and oncogenesis. Forkhead DNA- binding domain recognizes sequences containing the core motif 5'-(A/C)AA(C/T)A-3' with the binding stoichiometry 1:1. Results of this bachelor thesis show that FOXO4-DBD binds the...
Structural study of the ASK1:thioredoxin complex.
Pšenáková, Katarína
5 ABSTRACT The mitogen-activated protein kinase (MAPK) cascade is an essential member of the cell defense system against stressors. The capability and efficiency of the cell reactions to different stress signals depend on signal transduction pathway, where signals from MAPK kinase kinase (MAP3K) are transferred through phosphorylation to downstream MAPK kinase (MAP2K) and finally to MAPK. Apoptosis signal-regulating kinase 1 (ASK1) is a member of a MAP3K family and its activation and inhibition has a significant participation in a regulation of cell response to stress stimuli. The regulation of ASK1 has a strong influence in pathogenesis of several diseases, the excessive activation of human ASK1 or failure in the control of its function are associated with cardiovascular diseases, neurodegenerative disorders, inflammatory diseases, infectious diseases, tumorigenesis, asthma, diabetes and ageing. The activity of ASK1 is regulated by its interaction with several proteins, the attention is focused on two physiological inhibitors, mammalian thioredoxin (TRX) and the 14-3-3 protein. ASK1 in its inactive form is inhibited by bonds formation with TRX and 14-3-3, however the explicit mechanism of this interaction is unclear due to the absence of structural data. This work is a part of an extensive research about...
Structural studies of selected signaling protein complexes.
Pšenáková, Katarína ; Obšil, Tomáš (advisor) ; Hrabal, Richard (referee) ; Maloy Řezáčová, Pavlína (referee)
The ability of proteins to bind other molecules in response to various stimuli in their microenvironment serves as a platform for extensive regulatory networks coordinating downstream cell actions. The correct function of these signaling pathways depends mostly on noncovalent interactions often affecting the structure of proteins and protein complexes. Understanding the molecular mechanism of a protein function in cell signaling therefore often depends on our knowledge of a three-dimensional structure. In this doctoral thesis, I present the work that led to the understanding of several protein-protein and protein-ligand interactions implicated in cell signaling at the molecular level. I applied nuclear magnetic resonance spectroscopy, small angle X-ray scattering and other biophysical methods to determine the molecular basis of inhibition of four signaling proteins: Calcium/Calmodulin (Ca2+ /CaM)-dependent protein kinase kinase 2 (CaMKK2); protease Caspase-2; Forkhead transcription factor FOXO3, and Apoptosis signal-regulating protein kinase 1 (ASK1). In particular, I investigated the distinct roles of 14-3-3 and Ca2+ /CaM in the regulation of CaMKK2 activity. I also studied in detail the mechanism how 14-3-3 interferes with the caspase-2 oligomerization and its nuclear localization as well as...
Structural study of the ASK1:thioredoxin complex.
Pšenáková, Katarína
5 ABSTRACT The mitogen-activated protein kinase (MAPK) cascade is an essential member of the cell defense system against stressors. The capability and efficiency of the cell reactions to different stress signals depend on signal transduction pathway, where signals from MAPK kinase kinase (MAP3K) are transferred through phosphorylation to downstream MAPK kinase (MAP2K) and finally to MAPK. Apoptosis signal-regulating kinase 1 (ASK1) is a member of a MAP3K family and its activation and inhibition has a significant participation in a regulation of cell response to stress stimuli. The regulation of ASK1 has a strong influence in pathogenesis of several diseases, the excessive activation of human ASK1 or failure in the control of its function are associated with cardiovascular diseases, neurodegenerative disorders, inflammatory diseases, infectious diseases, tumorigenesis, asthma, diabetes and ageing. The activity of ASK1 is regulated by its interaction with several proteins, the attention is focused on two physiological inhibitors, mammalian thioredoxin (TRX) and the 14-3-3 protein. ASK1 in its inactive form is inhibited by bonds formation with TRX and 14-3-3, however the explicit mechanism of this interaction is unclear due to the absence of structural data. This work is a part of an extensive research about...
Structural study of the ASK1:thioredoxin complex.
Pšenáková, Katarína ; Obšil, Tomáš (advisor) ; Štěpánek, Miroslav (referee)
5 ABSTRACT The mitogen-activated protein kinase (MAPK) cascade is an essential member of the cell defense system against stressors. The capability and efficiency of the cell reactions to different stress signals depend on signal transduction pathway, where signals from MAPK kinase kinase (MAP3K) are transferred through phosphorylation to downstream MAPK kinase (MAP2K) and finally to MAPK. Apoptosis signal-regulating kinase 1 (ASK1) is a member of a MAP3K family and its activation and inhibition has a significant participation in a regulation of cell response to stress stimuli. The regulation of ASK1 has a strong influence in pathogenesis of several diseases, the excessive activation of human ASK1 or failure in the control of its function are associated with cardiovascular diseases, neurodegenerative disorders, inflammatory diseases, infectious diseases, tumorigenesis, asthma, diabetes and ageing. The activity of ASK1 is regulated by its interaction with several proteins, the attention is focused on two physiological inhibitors, mammalian thioredoxin (TRX) and the 14-3-3 protein. ASK1 in its inactive form is inhibited by bonds formation with TRX and 14-3-3, however the explicit mechanism of this interaction is unclear due to the absence of structural data. This work is a part of an extensive research about...
Study of intermolecular interactions by isothermal titration calorimetry
Pšenáková, Katarína ; Obšil, Tomáš (advisor) ; Alblová, Miroslava (referee)
Calorimetric methods are used to studying the mechanism of regulation and control of biological processes at the molecular level. Isothermal titration calorimetry (ITC) is used for monitoring the heat exchange of molecular interactions in an environment with constant temperature. This method is also preferred because of its ability of direct measurement of thermodynamic parameters associated with complex formation. ITC measures mostly the energetics of biochemical reactions or molecular interactions protein-protein, protein-ligand, or enzyme-substrate. Main goals of this bachelor thesis were: (I) to acquire working skills with nano isothermal titration calorimeter; (II) to prepare the DNA-binding domain of human transcription factor FOXO4 (FOXO4-DBD); and (III) to study thermodynamic aspects of the interaction between FOXO4-DBD and the double-stranded DNA containing specific binding motif. FOXO4 protein is one of the four members of a "O" subgroup of forkhead transcription factors FOX, which play an important role in many cellular processes including stress resistance, age control and oncogenesis. Forkhead DNA- binding domain recognizes sequences containing the core motif 5'-(A/C)AA(C/T)A-3' with the binding stoichiometry 1:1. Results of this bachelor thesis show that FOXO4-DBD binds the...

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