In collaboration with L. Krasný (Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic), we study subunits and sigma-factors unique for RNA polymerase of Gram-positive bacteria. We have determined structure of well-ordered N-terminal domain of delta subunit and characterized structural features of its intrinsically disordered C-terminal domain. In addition to the structural description, we characterized dynamics of both domains at various time scales using NMR relaxation. Investigation of other subunits and sigma-factors is in progress.
Multidisciplinary research of proteins involved in plant hormone signaling cascades is an example of a collaborative project within CEITEC (J. Hejtko, Functional Genomics and Proteomics of Plants). We have studied dynamics of receiver domain of the sensory kinase CKI1 and characterized its interactions with Mg(2+) ions and a phosphate analogue. We are involved in characterization of interactions of the CKI1 receiver domain with its down-stream partners and in structural studies of other sensory kinases (e.g. ETR1).
Traditionally, the strength of the group has been development of NMR methodology for atomic-resolution studies of proteins and nucleic acids. Recently, several fruitful collaborations witnessed that we are able to apply our know-how to solve real biological problems that were hard to attack by the conventional approaches. In the future, we would like to continue to develop in this direction and to complement the existing successful collaboration with our own projects. We plan to study dynamics, structural properties, function and regulation of Microtubule Associated Protein 2c (MAP2c), a cytoskeletal protein important for development of neuronal cells. The free form of the protein is disordered and relatively large (49 kDa), which makes it a challenging target for NMR. Our preliminary results show that the methodology developed in our group is sufficient for such a type of molecule.
3D structure of the delta subunit of RNA polymerase from Bacillus subtilis including disordered C-terminal tail determined by NMR spectroscopy.
list / cards
Name and position |
Phone |
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Lukáš Žídek, Ph.D. Research Group Leader |
+420 54949 8393 | |
Prof. Vladimír Sklenář Research Infrastructures Coordinator |
+420 54949 7022 | |
Jozef Hritz, Ph.D. Researcher |
+420 54949 3847 | |
Petr Padrta, Ph.D. Researcher |
+420 54949 6355 | |
Tanvir Shaikh Researcher |
+420 54949 7577 | |
Arnošt Mládek, Ph.D. Postdoctoral Fellow |
+420 54949 5398 | |
Zuzana Gelová PhD student |
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Kateřina Hanáková Research specialist - PhD student |
+420 54949 8442 | |
Dominika Ledvinová Research specialist - PhD student |
+420 54949 8442 | |
Zuzana Jaseňáková PhD student |
+420 54949 2615 | |
Kateřina Melková PhD student |
+420 54949 2523 | |
Vojtěch Zapletal PhD student |
+420 54949 8147, +420 54949 2523 | |
Šimon Džatko Student |
+420 54949 7832 | |
Martin Gajarský Student |
+420 54949 7832 | |
Michaela Krafčíková Student |
+420 54949 7832 | |
Olga Otrusinová PhD student |
+420 54949 2615 | |
Zuzana Trošanová odborná pracovnice - PhD student |
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Séverine Jansen Researcher |
+420 54949 2615 | |
Hana Konečná Specialist |
+420 54949 5050 | |
Tereza Chmelíková Student |
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Michaela Markvartová |
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Hana Štégnerová |
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Vladimír Jonas |
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Sudhir Kumar Pal |
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Jakub Šebera |
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Pavlína Víšková Student |
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Hana Zigová Student |
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Daniel Krafčík Student |
Supervisor: prof. Mgr. Lukáš Žídek, Ph.D.
Consultant: RNDr. Mgr. Jozef Hritz, Ph.D.
Microtubule Associated Protein 2c (MAP2c) is an intrinsically disordered protein important for development of nerve cells, expressed prenatally in dendrites. Dynamics, interactions, and structural features of MAP2c has been studied previously in our group. The aim of the thesis is to use nuclear magnetic resonance spectroscopy, X-ray crystallography, and other relevant techniques to characterize with atomic resolution structures of MAP2c present in complexes with interacting partners and transiently in solution.
Keywords: Microtubule Associated Protein 2c, MAP2c, NMR spectroscopy, X-ray crystallography, transient structures, nerve cells
Supervisor: RNDr. Mgr. Jozef Hritz, Ph.D.
Consultant: doc. Mgr. Vítězslav Bryja, Ph.D., Mgr. Jan Přibyl, Ph.D.
The neuronal cells of patients suffering from neurodegenerative diseases are characteristic by possession of aggregates of several proteins such as tau and Abeta protein. They form toxic aggregates to the different extend depending on the environment and their state (e.g. different post-translational modifications or specific conformational state). The main aim of this PhD project will be to quantify the extent of aggregation of three selected proteins in vitro and in the cell environment. Dependence of aggregation on phosphorylation and the potential interaction with various forms of 14-3-3 proteins will be analyzed mostly by biophysical interaction techniques and the atomic force microscopy. The biological relevance of the in-vitro findings will be tested in cell cultures by fluorescence microscopy.
Keywords: tau and Abeta protein, post-translational modifications, phosphorylation, 14-3-3 proteins, atomic force microscopy, neurodegenerative diseases, AFM
Supervisor: prof. Mgr. Lukáš Žídek, Ph.D.
Consultants: RNDr. Mgr. Jozef Hritz, Ph.D., doc. RNDr. Jan Hejátko, Ph.D.
The aim of the thesis is to study the role of conformational changes in function of the receiver domain of hybrid histidine kinase CKI1 from the model plant Arabidopsis thaliana and compare it with homologous histidine kinases from the same organism. Although the activation has been studied in structurally similar bacterial domains, little is known about the mechanism and structural basis of this event in plant systems, differing in several significant aspects. Our previous results show that the activation of the receiver domain is associated with a dramatic reduction of conformational dynamics of a loop in a vicinity of the phosphorylation site. NMR and computational approaches will be employed to study the structural changes during activation, to relate the dynamics of the loop to the function of other residues important for signal transduction, and to investigate interactions.
Keywords: hybrid histidine kinase, CKI1, Arabidopsis thaliana, NMR, conformational changes
Supervisor: RNDr. Mgr. Jozef Hritz, Ph.D.
Consultant: doc. Mgr. Vítězslav Bryja, Ph.D., Dr. Tanvir Shaikh
Several neurodegenerative diseases are asociated with the formation of fibrous (fibrillar) protein agreggates. The fibrillization of amyloid beta peptide into amyloid plaques and the agregation of hyperphosphorylated tau protein into neurofibrillar tangles are main neuropatological signs of Alzheimer disease. Studying of how different factors influence the formation of protein fibrils is the key for understanding this neurodegerative processes. The main aim of this PhD project will be characterization of structural changes in the formation of protein fibrils due to different phosphorylation state and the interaction with 14-3-3 proteins. Interdisciplinary approach combining molecular biology and structural biology (mainly cryoEM tomography) methods will be applied. Molecular binding mode involving 14-3-3 will be elucidated by NMR.
Keywords: neurodegenerative diseases, Alzheimer disease, protein fibrils, phosphorylation state, 14-3-3 proteins, cryoEM tomography, NMR
Supervisor: prof. Mgr. Lukáš Žídek, Ph.D.
Consultant: Mgr. Pavel Kadeřávek, Ph.D.
Certain RNA polymerase subunits (including sigma factors) are specific for Gram-positive bacteria. Such subunits are critical determinants of the enzyme specificity for DNA promoter sequence. Structures of delta subunits and of the vegetative sigmaA 1.1. domain, both containing highly dynamical regions, have been solved and further characterized in the group. The results suggest an important role of the conformational flexibility in functions of the subunits, but mechanistic details remain to be revealed. The aim of the thesis is to study dynamics and interactions of the subunit in order to understand their roles on the atomistic level, and provide structural data for other sigma factors and/or their domains. Standard and advanced methods of nuclear magnetic spectroscopy, recently developed in the group for such applications, will be combined with X-ray crystallography and cryo-electron microscopy
Keywords: Gram-positive bacteria, RNA polymerase subunits, NMR spectroscopy, X-ray crystallography, cryo-electron microscopy
29. ledna 2018 9:46
LECTURE: Dr. Ondrej Hovorka: Models of magnetic nanoparticles for biomedical applications
25. ledna 2018 18:21
WHEN: 30. 01. 2018 WHERE: CEITEC BUT, Purkynova 123, large meeting room SPEAKER: Dr Andriy Marko TALK: Advances in PELDOR…