Study of the interaction between DNA and transcription factors using mass spectrometry.

Transkripční faktory hrají v rámci buňky i celého mnohobuněčného organismu zásadní regulační úlohu. Důležitým faktorem, je jejich schopnost interagovat s ostatními regulačními proteiny a DNA. Přesto, že rozsáhlá část této interakční sítě je již zmapována, detailních informací o struktuře a dynamice protein-proteinových a protein-DNA komplexů je dosud velmi málo. V této práci jsme se zaměřili na možnosti studia konformačních změn daných tvorbou komplexu transkripčního faktoru s DNA pomocí strukturních metod hmotnostní spektrometrie: vodík/deuteriové výměny a chemického sítění. Jako modelový transkripční faktor jsme vybrali FOXO4, jehož DNA vazebná doména je strukturně dobře charakterizovaná jak ve volné formě, tak v komplexu s DNA (In Czech).Transcription factors play crucial regulatory role within the cell and the entire multicellular organism. The important factor is its ability to interact with other regulatory proteins and DNA. Despite the fact that a large part of the interaction network is already documented, detailed information on the structure and dynamics of protein-protein and protein-DNA complexes is still scarce. In this thesis we focused on the possibility of studying conformational changes given by the transcription factor-DNA complex formation using the methods of structural mass spectrometry: hydrogen/deuterium exchange and chemical crosslinking. As a model, we chose a transcription factor FOXO4 which DNA binding domain is structurally well characterized both in free form and in the complex with DNA.Department of BiochemistryKatedra biochemieFaculty of SciencePřírodovědecká fakult

Preparation of plasmids for expression of interleukin 2-fusion monoclonal antibody

Aim of the thesis was optimization of production of monoclonal antibody against interleu-kin 2 (IL 2) as fusion construct with IL 2 by preparation of new expression vectors based on pTW5 plasmid and by replacing the native signal sequence with new one, originally from secreted alkaline phosphatase. This fusion antibody has interesting biological activity with therapeutic potential - prolonging the half-life of IL 2 in blood circulation. (The thesis is written in Czech.) Powered by TCPDF (www.tcpdf.org

The process of Setting Up the Local Government in the Municipality of Benešov

This bachelor thesis focuses on local politics in Benešov. Thesis is separated into theoretic part and analytic part. Theoretic part contains definitions of basic terms and mechanisms that are needed to know to understand local politics. Analytic part is focused on Benešov and its demographic and socioeconomic characteristic, analysis of local elections results 2006, 2010 and 2014. Thesis includes also evaluation of actual political situation

MS-Based Approaches Enable the Structural Characterization of Transcription Factor/DNA Response Element Complex

The limited information available on the structure of complexes involving transcription factors and cognate DNA response elements represents a major obstacle in the quest to understand their mechanism of action at the molecular level. We implemented a concerted structural proteomics approach, which combined hydrogen-deuterium exchange (HDX), quantitative protein-protein and protein-nucleic acid cross-linking (XL), and homology analysis, to model the structure of the complex between the full-length DNA binding domain (DBD) of Forkhead box protein O4 (FOXO4) and its DNA binding element (DBE). The results confirmed that FOXO4-DBD assumes the characteristic forkhead topology shared by these types of transcription factors, but its binding mode differs significantly from those of other members of the family. The results showed that the binding interaction stabilized regions that were rather flexible and disordered in the unbound form. Surprisingly, the conformational effects were not limited only to the interface between bound components, but extended also to distal regions that may be essential to recruiting additional factors to the transcription machinery. In addition to providing valuable new insights into the binding mechanism, this project provided an excellent evaluation of the merits of structural proteomics approaches in the investigation of systems that are not directly amenable to traditional high-resolution techniques