Research
Focus
Structural mass spectrometry
method development
- novel covalent labeling and cross-linking chemistry of proteins /nucleic acids
- new acid proteases as a tool for protein digestion
- automation of HDX-MS and CX-MS workflow including software design
Functionalized surfaces for mass spectrometry
- ambient ion landing protein immobilization
- surface Immuno-affinity substrates for clinical diagnostics
- biochemically active plates for desorption mass spectrometry
Projects
- European Network of Fourier-Transform Ion-Cyclotron-Resonance Mass Spectrometry Centers
- Mapping the protein surface accessible area utilizing Top Down mass spectrometry and reactive radical footprinting.
- European Proteomics Infrastructure Consortium providing Access.
- Mass Spectrometric 3D Structure Analysis of DNA Response Elements / Transcription Factor Association and Modeling
- Structural and functional analysis of Hsp70/Hsp90 chaperone complexes
- A new tool for structure biology: Combination of high resolution mass spectrometry, chemical cross-linking and H/D exchange
- Structural mass spectrometry analysis for complexes of transcription factors with DNA response elements
- Electron Transfer in Cellulose Degrading Enzymes
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European Network of Fourier-Transform Ion-Cyclotron-Resonance Mass Spectrometry Centers
EU_FT-ICR_MS proposal aims to establish a European network of FT-ICR (Fourier Transform Ion Cyclotron Resonance) mass spectrometry (MS) centers in association with a manufacturer and a SME software company. Mass spectrometry (MS) has become the most ubiquitous analytical techniques in use today, providing more information on the composition and the structure of a substance from a smaller amount of sample than any other techniques. Unlike other analytical techniques, such as NMR, which mainly rely on a unique technology, MS is characterized by the existence of a large range of mass analyzers. FT-ICR MS is the most powerful MS technique. It offers up to 100 fold higher mass resolving power and mass accuracy than any other MS technique. On the contrary to NMR community with which the FT-ICR MS shares several features, FT-ICR MS has never been involved in a European INFRA network and so will be a legitimate candidate to the Integrating Activities for Starting Communities call. The EU_FT-ICR_MS network includes 10 FT-ICR MS centers from 8 different European countries (Belgium, Czech Republic, Finland, France, Germany, Italy, Portugal, and United Kingdom) and 1 third country (Russia), a European FT-ICR MS manufacturer and 2 SMEs. It includes centers equipped with up-to-date FT-ICR MS and expertise which will cover most of the field in which FT-ICR mass spectrometry is involved: BioOrganic & BioInorganic, Cultural heritage, Glycomics, Environment, Imaging, InfraRed Spectroscopy of Ions in the Gas Phase, Lipidomics, Medicine, Petroleum & Coal Oil, Nanoparticles, Organic chemistry, Physical chemistry, Proteomics, Structural biology. The EU_FT-ICR_MS proposal contains six work-packages which cover all the aspects of the INFRAIA-02-2017 (RIA) Integrating Activities for Starting Communities (WP1 Transnational access; WP2 Training and Education; WP3 Open Data and e-Infrastructure; WP 4 Joint Research Action; WP 5 Dissemination; WP6 Consortium management).
Funded by EU H2020 (EU_FT-ICR_MS)
Mapping the protein surface accessible area utilizing Top Down mass spectrometry and reactive radical footprinting.
Structural proteomics has become an emerging technique of current structural biology. It covers very broad range of analytical methods from hydrogen/deuterium exchange to chemical crosslinking. Although, the limitations and potentials of both techniques are well studied, foundations of covalent labelling are still poorly described. While H/D exchange offers information about backbone accessibility, covalent labelling targets amino acid side chains and thus provides complementary structural information. In this proposal we will develop chemical reagents with novel reactivity that will allow targeting of more amino acid side chains and expand the spatial resolution of the method. We will also pursue hydroxyradical footprinting where we aim mainly at the development of top-down mass spectrometric approaches allowing to assign not only the site of the modification but also to assess the reactivity scale of individual modified residues. Such advancements will greatly facilitate protein structure characterization and will provide valuable information for protein structure modelling.
Funded by Czech Science Foundation (19-16084S)
European Proteomics Infrastructure Consortium providing Access.
In life science research, proteomics technologies promise to be even more revolutionary than genomics for understanding the molecular mechanisms of complex biological systems and their (mal-)functions in health and disease. As a result, there is a strongly growing need for the integration of proteomic technologies into world-class life science research programs in medicine, agriculture, food, biotechnology, and materials. The European Proteomics Infrastructure Consortium providing Access (EPIC-XS) brings together a consortium of worldleading, highly innovative European proteomics facilities with the shared goal of supporting excellent life science research in Europe. This will be achieved by providing access to cutting-edge proteomics facilities, and by developing and implementing novel mass spectrometry based proteomics and bioinformatics approaches to shape the future life science research in Europe. The Transnational Access in EPIC-XS consists of around 2400 days of access to state-of-the-art proteomics facilities, corresponding to around 150 user projects, which will be evaluated by an independent review panel to ensure excellence, and to match users with the best facility for their challenging research questions. The cutting-edge Joint Research Activities in EPIC-XS are built on the extremely strong innovation track record of the consortium members, and will address bottlenecks in, and develop novel approaches for, future-oriented efforts in computational and structural proteomics, analysis of higher-order proteome organization, multi-omics integration, and translational/clinical proteomics and personalized/precision medicine. EPIC-XS will provide users with unrivalled access to world-leading facilities and expertise, while simultaneously extending Europe’s leading international role in the. EPIC-XS meets the widespread current demands for user access, addresses present bottlenecks in the field, and will open up entirely new perspectives in proteomics.
Funded by EU H2020 (EPIC-XS)
Mass Spectrometric 3D Structure Analysis of DNA Response Elements / Transcription Factor Association and Modeling
The structural analysis of heterogeneous native biomolecular interactions in vivo is hampered by the lack of suitable fast and high-throughput methods with an appropriate resolving power. The 2D proteomic mass spectrometric (MS)-based analysis of the interactome currently represents the gold standard for the target identification and novel drug screening. A cohort of new structural mass spectrometric methods (MS3D) supported by molecular dynamics (MD) modeling algorithms is fully complementary to the existing high-resolution but non-native structural methods. The goal of this project is to develop the MS3D method of biomolecular interaction pairs of the transcription factor (TF) and its DNA response element, both in vitro and in vivo models. The low-resolution analysis of the individual structures and their interaction landscapes will deliver experimental data feedback to the predictive MD computation of native heterogeneous complexes. The ultimate mission is to establish the MS3D native structural analysis as a novel standard of biomolecular analysis for the rational drug design.
Funded by Czech Science Foundation (16-24309S)
Structural and functional analysis of Hsp70/Hsp90 chaperone complexes
Project is focused on the analysis of interactions between chaperones Hsp90 and Hsp70 and their co-chaperones. Combination of structural mass spectrometry with functional assays will be used to describe the interactions sites as well as parts of the molecules influenced by the formation of chaperone complexes. This approach should reveal new factors influencing the activity of Hsp70 and Hsp90, uncover new protein interaction sites suitable for designing new low molecular weight inhibitors and bring insights into the dynamic behaviour of the chaperone complexes. The new findings related to assembly and functioning of chaperone complexes can also uncover the mechanisms that underlie several human diseases including cancer or neurodegenerative diseases. Besides the contribution to basic research, the results of the project can be exploited in biotechnology industry including rational drug design or folding of recombinant proteins.
Joint project with Dr. Petr Muller, RECAMO, Brno, Czech Republic
Funded by Czech Science Foundation (16-20860S)
A new tool for structure biology: Combination of high resolution mass spectrometry, chemical cross-linking and H/D exchange
The aim of this project is to nucleate a group of researchers with a common interest, namely developing and applying new biomolecular mass spectrometry (MS) methods in order to make the characterisation of protein structure and dynamcis more rapid and routine. Methods include non-denaturing MS approaches in combination with ion mobility, as well as hydrogen-deuterium exchange, chemical crosslinking and other labeling techniques together with computational approaches. This toolbox will be made available to the broader scientific community, and will greatly enhance our ability to design new drugs and ensure the quality and efficacy of biopharmaceuticals, thereby benefiting human health.
Funded by Ministry of Education, Youth, and Sports (COST CZ – LH15010)
Structural mass spectrometry analysis for complexes of transcription factors with DNA response elements
The proposed project is aimed to in-vitro and in-vivo identification of DNA response elements using chemical cross-linking and hydrogen/deuterium exchange combined with high resolution mass spectrometry. The next goal is participation of young scientists in the international frame of this research project.
New knowledge will lead not only to valuable insight into regulation transcription and gene expression, but also into the structure-function relationship of complexes transcription factor/DNA.
Joint project with Prof Daniele Fabris, State University of New York, Albany, U.S.A.
Funded by Ministry of Education, Youth, and Sports (Kontakt II – LH15010)
Electron Transfer in Cellulose Degrading Enzymes
The proposed project is aimed at elucidation of interactions between cellobiose dehydrogenase (CDH) and its putative redox partner – lytic polysaccharide monooxygenase (LPMO) – in solution and in their cellulose bound state. Our research strategy is based on the combined expertise of both research partners. The Austrian group will perform protein engineering and production as well as in-solution spectrophotometry and calorimetry to monitor CDH-LPMO electron transfer rates and interactions. The Czech team will employ structural mass spectrometry techniques based on protein isotopic exchange and chemical cross-linking to investigate the structural details of the protein-protein and protein-substrate interactions. The results will answer the questions if CDH functions as an “LPMO reductase” and if specific interactions are involved in their interprotein electron transfer. This will expand our understanding of biocatalytic redox processes of cellulose depolymerization, which may help increasing the efficiency of biofuel production or degradation processes in biorefineries.
Collaboration with Petr Halada - joint project with prof. Roland Ludwig, University of Natural Resources and Life Sciences, Vienna, Austria
Funded by Czech Science Foundation and Fonds zur Förderung der wissenschaftlichen Forschung (Austira) – (I 2385-N28)