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Molecular mechanisms of signal transduction in model heme-containing oxygen sensor proteins
Stráňava, Martin ; Martínková, Markéta (advisor) ; Obšil, Tomáš (referee) ; Macek, Tomáš (referee)
EN Heme containing gas sensor proteins play important role in bacterial physiology in regulating many processes such as cell differentiation, virulence, biofilm formation or intercellular communication. For their structure, typical modular architecture is characteristic where various sensor domains (usually at the N-terminus) regulate the activity of the catalytic or functional domains (usually at the C-terminus). In this dissertation thesis, we focused on three representatives from the group of oxygen sensing proteins, namely histidine kinase AfGcHK, diguanylate cyclase YddV, phosphodiesterase EcDOS and also on protein RR, which is the interaction partner of AfGcHK. The main aim of the thesis was to study intra-protein/inter-domain signal transduction in two representatives of heme sensor proteins with a globin fold of the sensor domain (AfGcHK, YddV) and in one representative with PAS fold of the sensor domain (EcDOS). Another objective was to describe inter-protein signal transduction in the two component signaling system AfGcHK-RR and structurally characterize these two interacting partners. Emphasis was also placed on the study of the interaction between model sensor domains and different signaling molecules and also on function of individual amino acids involved in the binding of these...
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Enzyme activity analysis of function domains belonging to model heme-containing sensor proteins
Prošková, Veronika
EN This Ph.D. thesis focuses on the heme containing gas sensor proteins. These proteins are predominantly present in bacteria, in which play an important role in processes like, sporulation, antibiotic resistance and so on. Heme containing sensor proteins composed of two domains. First one is a globin domain, which contains the heme molecule. Interaction of heme with gas molecule acts as a signal for the activation/inactivation of the second functional domain. Part of this thesis is formed by a review, which summarized the current knowledge about heme containing sensor proteins. In the next part of this thesis we focused on three representatives from the group of oxygen sensor proteins - histidine kinase AfGcHK, diguanylate cyclase YddV and phosphodiesterase EcDOS. The main aim of this thesis was to solve the mechanism of interdomain/intraprotein signal transduction in two oxygen sensor proteins with globin fold of their sensor domain (AfGcHK, YddV). For this purpose, we used the kinetic analysis of their functional domain activity and the methods of structural biology. We also studied the mechanism of interprotein signal transduction in AfGcHK and its cognate partner RR protein. It was also tested, how the presence of sodium disulfide affects the functional properties of oxygen sensor proteins...
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Enzyme activity analysis of function domains belonging to model heme-containing sensor proteins
Prošková, Veronika
EN This Ph.D. thesis focuses on the heme containing gas sensor proteins. These proteins are predominantly present in bacteria, in which play an important role in processes like, sporulation, antibiotic resistance and so on. Heme containing sensor proteins composed of two domains. First one is a globin domain, which contains the heme molecule. Interaction of heme with gas molecule acts as a signal for the activation/inactivation of the second functional domain. Part of this thesis is formed by a review, which summarized the current knowledge about heme containing sensor proteins. In the next part of this thesis we focused on three representatives from the group of oxygen sensor proteins - histidine kinase AfGcHK, diguanylate cyclase YddV and phosphodiesterase EcDOS. The main aim of this thesis was to solve the mechanism of interdomain/intraprotein signal transduction in two oxygen sensor proteins with globin fold of their sensor domain (AfGcHK, YddV). For this purpose, we used the kinetic analysis of their functional domain activity and the methods of structural biology. We also studied the mechanism of interprotein signal transduction in AfGcHK and its cognate partner RR protein. It was also tested, how the presence of sodium disulfide affects the functional properties of oxygen sensor proteins...
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Enzyme activity analysis of function domains belonging to model heme-containing sensor proteins
Prošková, Veronika ; Martínková, Markéta (advisor) ; Macek, Tomáš (referee) ; Vítek, Libor (referee)
EN This Ph.D. thesis focuses on the heme containing gas sensor proteins. These proteins are predominantly present in bacteria, in which play an important role in processes like, sporulation, antibiotic resistance and so on. Heme containing sensor proteins composed of two domains. First one is a globin domain, which contains the heme molecule. Interaction of heme with gas molecule acts as a signal for the activation/inactivation of the second functional domain. Part of this thesis is formed by a review, which summarized the current knowledge about heme containing sensor proteins. In the next part of this thesis we focused on three representatives from the group of oxygen sensor proteins - histidine kinase AfGcHK, diguanylate cyclase YddV and phosphodiesterase EcDOS. The main aim of this thesis was to solve the mechanism of interdomain/intraprotein signal transduction in two oxygen sensor proteins with globin fold of their sensor domain (AfGcHK, YddV). For this purpose, we used the kinetic analysis of their functional domain activity and the methods of structural biology. We also studied the mechanism of interprotein signal transduction in AfGcHK and its cognate partner RR protein. It was also tested, how the presence of sodium disulfide affects the functional properties of oxygen sensor proteins...
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Molecular mechanisms of signal transduction in model heme-containing oxygen sensor proteins
Stráňava, Martin ; Martínková, Markéta (advisor) ; Obšil, Tomáš (referee) ; Macek, Tomáš (referee)
EN Heme containing gas sensor proteins play important role in bacterial physiology in regulating many processes such as cell differentiation, virulence, biofilm formation or intercellular communication. For their structure, typical modular architecture is characteristic where various sensor domains (usually at the N-terminus) regulate the activity of the catalytic or functional domains (usually at the C-terminus). In this dissertation thesis, we focused on three representatives from the group of oxygen sensing proteins, namely histidine kinase AfGcHK, diguanylate cyclase YddV, phosphodiesterase EcDOS and also on protein RR, which is the interaction partner of AfGcHK. The main aim of the thesis was to study intra-protein/inter-domain signal transduction in two representatives of heme sensor proteins with a globin fold of the sensor domain (AfGcHK, YddV) and in one representative with PAS fold of the sensor domain (EcDOS). Another objective was to describe inter-protein signal transduction in the two component signaling system AfGcHK-RR and structurally characterize these two interacting partners. Emphasis was also placed on the study of the interaction between model sensor domains and different signaling molecules and also on function of individual amino acids involved in the binding of these...
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Effect of sodium sulfide on the propreties of model globine-coupled heme-containing sensor proteins
Bartošová, Martina ; Martínková, Markéta (advisor) ; Man, Petr (referee)
Hydrogen sulfide mediates various physiological functions and along with carbon monoxide and nitric oxide it is an important gaseous signaling molecule. Cellular targets for H2S are proteins, enzymes, transcriptional factors or ion channels. In many cases, the effect of H2S on the regulatory protein is mediated by modifications of its cystein residues. In hemeproteins, the regulation of catalytic activity is induced by formation of the Fe(III)-SH complex or by reduction of the heme iron with subsequent formation of Fe(II)-O2 complex. The effect of Na2S on model sensor heme-containing proteins is presented in this thesis. Protein, isolated from bacterium Anaeromyxobacter sp. strain FW109-5, containing a globine coupled sensor domain and a histidine kinase domain is one of the studied proteins, the second one is protein isolated from bacterium Escherichie coli, containing a globine coupled sensor domain and a diguanylate cyclase domain. The effect of Na2S on both model proteins and their mutants was studied by UV-Vis spectral analysis. Spectra of YddV-HD Y43A were very unique, because thery confirmed formation of a homogenous complex Fe(III)-SH in this protein, whereas only mixtures of varous heme complexes were detected in other tested proteins. Additionally the effect of Na2S on functional domain...
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