19 results on '"Stefan Mogk"'
Search Results
2. Protocol to analyze the transmigration efficiency of T. brucei using an in vitro model of the blood-cerebrospinal fluid barrier
- Author
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Alexander Herrmann, Carolin Susanne Schnedermann, Hiroshi Ishikawa, Christian Schwerk, Horst Schroten, and Stefan Mogk
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Cell biology ,Cell culture ,Cell-based assays ,Health sciences ,Microbiology ,Model organisms ,Science (General) ,Q1-390 - Abstract
Summary: At present, the only approach to investigate the transmigration of Trypanosoma brucei, the causative agent of human African trypanosomiasis, from blood to cerebrospinal fluid is through animal experiments. This protocol details how to analyze the transmigration efficiency using an in vitro model of the blood-cerebrospinal fluid (blood-CSF) barrier. We describe how to grow human choroid plexus epithelial cells on cell culture filter inserts to form the barrier, followed by isolating and quantifying genomic DNA of transmigrated parasites by qPCR.For complete details on the use and execution of this protocol, please refer to Speidel et al. (2022). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
- Published
- 2022
- Full Text
- View/download PDF
3. Transmigration of Trypanosoma brucei across an in vitro blood-cerebrospinal fluid barrier
- Author
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Annika Speidel, Marianne Theile, Lena Pfeiffer, Alexander Herrmann, Katherine Figarella, Hiroshi Ishikawa, Christian Schwerk, Horst Schroten, Michael Duszenko, and Stefan Mogk
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Microbiology ,Parasitology ,Bioengineering ,Science - Abstract
Summary: Trypanosoma brucei is the causative agent of human African trypanosomiasis. The parasite transmigrates from blood vessels across the choroid plexus epithelium to enter the central nervous system, a process that leads to the manifestation of second stage sleeping sickness. Using an in vitro model of the blood-cerebrospinal fluid barrier, we investigated the mechanism of the transmigration process. For this, a monolayer of human choroid plexus papilloma cells was cultivated on a permeable membrane that mimics the basal lamina underlying the choroid plexus epithelial cells. Plexus cells polarize and interconnect forming tight junctions. Deploying different T. brucei brucei strains, we observed that geometry and motility are important for tissue invasion. Using fluorescent microscopy, the parasite’s moving was visualized between plexus epithelial cells. The presented model provides a simple tool to screen trypanosome libraries for their ability to infect cerebrospinal fluid or to test the impact of chemical substances on transmigration.
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- 2022
- Full Text
- View/download PDF
4. Serial crystallography on in vivo grown microcrystals using synchrotron radiation
- Author
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Cornelius Gati, Gleb Bourenkov, Marco Klinge, Dirk Rehders, Francesco Stellato, Dominik Oberthür, Oleksandr Yefanov, Benjamin P. Sommer, Stefan Mogk, Michael Duszenko, Christian Betzel, Thomas R. Schneider, Henry N. Chapman, and Lars Redecke
- Subjects
protein microcrystallography ,serial crystallography ,in vivo grown microcrystals ,Crystallography ,QD901-999 - Abstract
Crystal structure determinations of biological macromolecules are limited by the availability of sufficiently sized crystals and by the fact that crystal quality deteriorates during data collection owing to radiation damage. Exploiting a micrometre-sized X-ray beam, high-precision diffractometry and shutterless data acquisition with a pixel-array detector, a strategy for collecting data from many micrometre-sized crystals presented to an X-ray beam in a vitrified suspension is demonstrated. By combining diffraction data from 80 Trypanosoma brucei procathepsin B crystals with an average volume of 9 µm3, a complete data set to 3.0 Å resolution has been assembled. The data allowed the refinement of a structural model that is consistent with that previously obtained using free-electron laser radiation, providing mutual validation. Further improvements of the serial synchrotron crystallography technique and its combination with serial femtosecond crystallography are discussed that may allow the determination of high-resolution structures of micrometre-sized crystals.
- Published
- 2014
- Full Text
- View/download PDF
5. Cyclical appearance of African trypanosomes in the cerebrospinal fluid: new insights in how trypanosomes enter the CNS.
- Author
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Stefan Mogk, Andreas Meiwes, Swetlana Shtopel, Ulrich Schraermeyer, Michael Lazarus, Bruno Kubata, Hartwig Wolburg, and Michael Duszenko
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Medicine ,Science - Abstract
It is textbook knowledge that human infective forms of Trypanosoma brucei, the causative agent of sleeping sickness, enter the brain across the blood-brain barrier after an initial phase of weeks (rhodesiense) or months (gambiense) in blood. Based on our results using an animal model, both statements seem questionable. As we and others have shown, the first infection relevant crossing of the blood brain border occurs via the choroid plexus, i.e. via the blood-CSF barrier. In addition, counting trypanosomes in blood-free CSF obtained by an atlanto-occipital access revealed a cyclical infection in CSF that was directly correlated to the trypanosome density in blood infection. We also obtained conclusive evidence of organ infiltration, since parasites were detected in tissues outside the blood vessels in heart, spleen, liver, eye, testis, epididymis, and especially between the cell layers of the pia mater including the Virchow-Robin space. Interestingly, in all organs except pia mater, heart and testis, trypanosomes showed either a more or less degraded appearance of cell integrity by loss of the surface coat (VSG), loss of the microtubular cytoskeleton and loss of the intracellular content, or where taken up by phagocytes and degraded intracellularly within lysosomes. This is also true for trypanosomes placed intrathecally into the brain parenchyma using a stereotactic device. We propose a different model of brain infection that is in accordance with our observations and with well-established facts about the development of sleeping sickness.
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- 2014
- Full Text
- View/download PDF
6. Late stage infection in sleeping sickness.
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Hartwig Wolburg, Stefan Mogk, Sven Acker, Claudia Frey, Monika Meinert, Caroline Schönfeld, Michael Lazarus, Yoshihiro Urade, Bruno Kilunga Kubata, and Michael Duszenko
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Medicine ,Science - Abstract
At the turn of the 19(th) century, trypanosomes were identified as the causative agent of sleeping sickness and their presence within the cerebrospinal fluid of late stage sleeping sickness patients was described. However, no definitive proof of how the parasites reach the brain has been presented so far. Analyzing electron micrographs prepared from rodent brains more than 20 days after infection, we present here conclusive evidence that the parasites first enter the brain via the choroid plexus from where they penetrate the epithelial cell layer to reach the ventricular system. Adversely, no trypanosomes were observed within the parenchyma outside blood vessels. We also show that brain infection depends on the formation of long slender trypanosomes and that the cerebrospinal fluid as well as the stroma of the choroid plexus is a hostile environment for the survival of trypanosomes, which enter the pial space including the Virchow-Robin space via the subarachnoid space to escape degradation. Our data suggest that trypanosomes do not intend to colonize the brain but reside near or within the glia limitans, from where they can re-populate blood vessels and disrupt the sleep wake cycles.
- Published
- 2012
- Full Text
- View/download PDF
7. Transmigration of
- Author
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Annika, Speidel, Marianne, Theile, Lena, Pfeiffer, Alexander, Herrmann, Katherine, Figarella, Hiroshi, Ishikawa, Christian, Schwerk, Horst, Schroten, Michael, Duszenko, and Stefan, Mogk
- Published
- 2021
8. The conserved hypothetical protein Tb427.10.13790 is required for cytokinesis in Trypanosoma brucei
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Salesia Franziska Werner, Christian Betzel, Michael Duszenko, Celestin Nzanzu Mudogo, and Stefan Mogk
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0301 basic medicine ,biology ,Veterinary (miscellaneous) ,Trypanosoma brucei brucei ,Hypothetical protein ,Protozoan Proteins ,Flagellum ,Trypanosoma brucei ,biology.organism_classification ,Cell biology ,03 medical and health sciences ,030104 developmental biology ,Infectious Diseases ,RNA interference ,Insect Science ,Kinetoplast ,parasitic diseases ,Antigenic variation ,Animals ,Parasitology ,Gene ,Cytokinesis - Abstract
Trypanosoma brucei, a flagellated protozoan causing the deadly tropical disease Human African Trypanosomiasis (HAT), affects people in sub-Saharan Africa. HAT therapy relies upon drugs which use is limited by toxicity and rigorous treatment regimes, while development of vaccines remains elusive, due to the effectiveness of the parasite´s antigenic variation. Here, we evaluate a hypothetical protein Tb427.10.13790, as a potential drug target. This protein is conserved among all kinetoplastids, but lacks homologs in all other pro- and eukaryotes. Knockdown of Tb427.10.13790 resulted in appearance of monster cells containing multiple nuclei and multiple flagella, a considerable enlargement of the flagellar pocket and eventually a lethal phenotype. Furthermore, analysis of kinetoplast and nucleus division in the knockdown cell line revealed a partial cell cycle arrest and failure to initiate cytokinesis. Likewise, overexpression of the respective protein fused with enhanced green fluorescent protein was also lethal for T. brucei. In these cells, the labelled protein appeared as a single dot near kinetoplast and flagellar pocket. Our results reveal that Tb427.10.13790 is essential for the parasite´s viability and may be a suitable new anti-trypanosomatid drug target candidate. Furthermore, we suggest that it might be worthwhile to investigate also other of the many so far just annotated trypanosome genes as a considerable number of them to lack human homologs but may be of critical importance for the kinetoplastid parasites.
- Published
- 2018
9. A Simple Approach to Perform TEER Measurements Using a Self-Made Volt-Amperemeter with Programmable Output Frequency
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Marianne Theile, Linus Wiora, Hiroshi Ishikawa, Dominik Russ, Stefan Mogk, Jonas Reuter, Christian Schwerk, and Horst Schroten
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General Immunology and Microbiology ,General Chemical Engineering ,General Neuroscience ,Cytological Techniques ,010401 analytical chemistry ,Endothelial Cells ,Volt ,Epithelial Cells ,Membrane filter ,02 engineering and technology ,Lower compartment ,021001 nanoscience & nanotechnology ,01 natural sciences ,General Biochemistry, Genetics and Molecular Biology ,Cell Line ,0104 chemical sciences ,Capillary Permeability ,Electrical resistance and conductance ,Electrode ,Electric Impedance ,Humans ,0210 nano-technology ,Electrodes ,Electrical impedance ,Biomedical engineering - Abstract
Transepithelial/endothelial electrical resistance (TEER) has been used since the 1980s to determine confluency and permeability of in vitro barrier model systems. In most cases, chopstick electrodes are used to determine the electric impedance between the upper and lower compartment of a cell culture filter insert system containing cellular monolayers. The filter membrane allows the cells to adhere, polarize, and interact by building tight junctions. This technique has been described with a variety of different cell lines (e.g., cells of the blood-brain barrier, blood-cerebrospinal fluid barrier, or gastrointestinal and pulmonary tract). TEER measurement devices can be readily obtained from different laboratory equipment suppliers. However, there are more cost-effective and customizable solutions imaginable if an appropriate voltammeter is self-assembled. The overall aim of this publication is to set up a reliable device with programmable output frequency that can be used with commercially available chopstick electrodes for TEER measurement.
- Published
- 2019
10. African trypanosomes and brain infection - the unsolved question
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Jasmin Stein, Celestin Nzanzu Mudogo, Christian Boßelmann, Stefan Mogk, Hartwig Wolburg, and Michael Duszenko
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0301 basic medicine ,biology ,Tsetse fly ,Trypanosoma brucei ,biology.organism_classification ,medicine.disease ,Virology ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,Cerebrospinal fluid ,Lymphatic system ,Immune system ,Immunology ,medicine ,Choroid plexus ,Glymphatic system ,General Agricultural and Biological Sciences ,Trypanosomiasis ,030217 neurology & neurosurgery - Abstract
African trypanosomes induce sleeping sickness. The parasites are transmitted during the blood meal of a tsetse fly and appear primarily in blood and lymph vessels, before they enter the central nervous system. During the latter stage, trypanosomes induce a deregulation of sleep-wake cycles and some additional neurological disorders. Historically, it was assumed that trypanosomes cross the blood-brain barrier and settle somewhere between the brain cells. The brain, however, is a strictly controlled and immune-privileged area that is completely surrounded by a dense barrier that covers the blood vessels: this is the blood-brain barrier. It is known that some immune cells are able to cross this barrier, but this requires a sophisticated mechanism and highly specific cell-cell interactions that have not been observed for trypanosomes within the mammalian host. Interestingly, trypanosomes injected directly into the brain parenchyma did not induce an infection. Likewise, after an intraperitoneal infection of rats, Trypanosoma brucei brucei was not observed within the brain, but appeared readily within the cerebrospinal fluid (CSF) and the meninges. Therefore, the parasite did not cross the blood-brain barrier, but the blood-CSF barrier, which is formed by the choroid plexus, i.e. the part of the ventricles where CSF is produced from blood. While there is no question that trypanosomes are able to invade the brain to induce a deadly encephalopathy, controversy exists about the pathway involved. This review lists experimental results that support crossing of the blood-brain barrier and of the blood-CSF barrier and discuss the implications that either pathway would have on infection progress and on the survival strategy of the parasite. For reasons discussed below, we prefer the latter pathway and suggest the existence of an additional distinct meningeal stage, from which trypanosomes could invade the brain via the Virchow-Robin space thereby bypassing the blood-brain barrier. We also consider healthy carriers, i.e. people living symptomless with the disease for up to several decades, and discuss implications the proposed meningeal stage would have for new anti-trypanosomal drug development. Considering the re-infection of blood, a process called relapse, we discuss the likely involvement of the newly described glymphatic connection between the meningeal space and the lymphatic system, that seems also be important for other infectious diseases.
- Published
- 2016
11. Life and Death of Trypanosoma Brucei : New Perspectives for Drug Development
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Michael Duszenko, Bruno Kilunga Kubata, Caroline Schönfeld, Torsten Barth, Stefan Mogk, and Jasmin Stein
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Programmed cell death ,Necrosis ,biology ,Autophagy ,Trypanosoma brucei ,medicine.disease ,biology.organism_classification ,Microbial Physiology ,Drug development ,Apoptosis ,Immunology ,medicine ,medicine.symptom ,Trypanosomiasis - Published
- 2013
12. African trypanosomes and brain infection - the unsolved question
- Author
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Stefan, Mogk, Christian M, Boßelmann, Celestin N, Mudogo, Jasmin, Stein, Hartwig, Wolburg, and Michael, Duszenko
- Subjects
Central Nervous System ,Trypanosomiasis, African ,Blood-Brain Barrier ,Animals ,Brain ,Humans ,Host-Parasite Interactions - Abstract
African trypanosomes induce sleeping sickness. The parasites are transmitted during the blood meal of a tsetse fly and appear primarily in blood and lymph vessels, before they enter the central nervous system. During the latter stage, trypanosomes induce a deregulation of sleep-wake cycles and some additional neurological disorders. Historically, it was assumed that trypanosomes cross the blood-brain barrier and settle somewhere between the brain cells. The brain, however, is a strictly controlled and immune-privileged area that is completely surrounded by a dense barrier that covers the blood vessels: this is the blood-brain barrier. It is known that some immune cells are able to cross this barrier, but this requires a sophisticated mechanism and highly specific cell-cell interactions that have not been observed for trypanosomes within the mammalian host. Interestingly, trypanosomes injected directly into the brain parenchyma did not induce an infection. Likewise, after an intraperitoneal infection of rats, Trypanosoma brucei brucei was not observed within the brain, but appeared readily within the cerebrospinal fluid (CSF) and the meninges. Therefore, the parasite did not cross the blood-brain barrier, but the blood-CSF barrier, which is formed by the choroid plexus, i.e. the part of the ventricles where CSF is produced from blood. While there is no question that trypanosomes are able to invade the brain to induce a deadly encephalopathy, controversy exists about the pathway involved. This review lists experimental results that support crossing of the blood-brain barrier and of the blood-CSF barrier and discuss the implications that either pathway would have on infection progress and on the survival strategy of the parasite. For reasons discussed below, we prefer the latter pathway and suggest the existence of an additional distinct meningeal stage, from which trypanosomes could invade the brain via the Virchow-Robin space thereby bypassing the blood-brain barrier. We also consider healthy carriers, i.e. people living symptomless with the disease for up to several decades, and discuss implications the proposed meningeal stage would have for new anti-trypanosomal drug development. Considering the re-infection of blood, a process called relapse, we discuss the likely involvement of the newly described glymphatic connection between the meningeal space and the lymphatic system, that seems also be important for other infectious diseases.
- Published
- 2016
13. Competence-based education in the example of a biochemical practical course with clinically relevant content for undergraduate medical students in the fourth semester of the preclinical section
- Author
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Stefan Mogk
- Subjects
Medical education ,Psychology ,Competence (human resources) - Abstract
This article was migrated. The article was not marked as recommended. Focus in traditional practical courses has always been on imparting technical knowledge. However, significance and application often are unclear for students, leading to low motivation. Thus, a practical course that is clinically relevant to students in the 4th semester was set up in biochemistry so that: a) a clear link to the clinical work can be identified, b) the roles of the physician, as set out in the German National Competence-based Learning Objectives Catalogue for Undergraduate Medical Education (NKLM), are covered in the best possible manner, and c) the scientific basis has a high importance.The newly established course covers key points of research work in 4 days, beginning with a fictitious case study. Samples of (artificial) urine and blood are used to diagnose sickle cell anemia and malaria, and their molecular basis is analysed using bioinformatic tools. Students learn how to research and discuss academic publications. Working in groups, they establish a concept using gene therapy to explore how a higher expression of fetal hemoglobin in erythroblasts could be induced using RNA interference to attenuate progression of the disease. Instead of recording the findings in a traditional manner, the students work together in groups to produce a manuscript of their results, which is critically examined to assess the suitability for being published.The semester cohort was randomly allocated to the newly established (n=79) and the traditional (n=57) practical course. Both courses were comparatively evaluated. The newly designed course received an overall assessment of 1.4, which was significantly better than the traditional course which received an overall assessment of 2.9.During the next years, nearly all subjects of the preclinical section will be challenged to find ways of combining NLKM requirements and technical knowledge. The concept presented here may serve as a model to be readily adapted to other fields.
- Published
- 2018
14. In vivo protein crystallization in combination with highly brilliant radiation sources offers novel opportunities for the structural analysis of post-translationally modified eukaryotic proteins
- Author
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Lars Redecke, Stefan Mogk, Dominik Oberthuer, Christian Betzel, Celestin Nzanzu Mudogo, B.P. Sommer, and Michael Duszenko
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Biophysics ,Gene Expression ,Nanotechnology ,Electrons ,CHO Cells ,Saccharomyces cerevisiae ,Biology ,Spodoptera ,Biochemistry ,law.invention ,Cathepsin B ,fluids and secretions ,Cricetulus ,Structural Biology ,law ,In vivo ,parasitic diseases ,Genetics ,Escherichia coli ,Sf9 Cells ,Animals ,Humans ,Crystallization ,Insect cell ,Crystallography ,Lasers ,computer.file_format ,Condensed Matter Physics ,Protein Data Bank ,equipment and supplies ,HEK293 Cells ,IYCr crystallization series ,Membrane protein ,Protein crystallization ,computer ,Protein Processing, Post-Translational ,Synchrotrons - Abstract
During the last decade, the number of three-dimensional structures solved by X-ray crystallography has increased dramatically. By 2014, it had crossed the landmark of 100 000 biomolecular structures deposited in the Protein Data Bank. This tremendous increase in successfully crystallized proteins is primarily owing to improvements in cloning strategies, the automation of the crystallization process and new innovative approaches to monitor crystallization. However, these improvements are mainly restricted to soluble proteins, while the crystallization and structural analysis of membrane proteins or proteins that undergo major post-translational modifications remains challenging. In addition, the need for relatively large crystals for conventional X-ray crystallography usually prevents the analysis of dynamic processes within cells. Thus, the advent of high-brilliance synchrotron and X-ray free-electron laser (XFEL) sources and the establishment of serial crystallography (SFX) have opened new avenues in structural analysis using crystals that were formerly unusable. The successful structure elucidation of cathepsin B, accomplished by the use of microcrystals obtained byin vivocrystallization in baculovirus-infected Sf9 insect cells, clearly proved that crystals grown intracellularly are very well suited for X-ray analysis. Here, methods by whichin vivocrystals can be obtained, isolated and used for structural analysis by novel highly brilliant XFEL and synchrotron-radiation sources are summarized and discussed.
- Published
- 2015
15. Brain infection by African trypanosomes during sleeping sickness
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Michael Duszenko, Hartwig Wolburg, Stefan Mogk, Bruno Kilunga Kubata, and Claudia Frey
- Subjects
Psychiatry and Mental health ,Brain infection ,business.industry ,General Neuroscience ,Immunology ,Medicine ,Neurology (clinical) ,business - Published
- 2012
16. The lane to the brain: how African trypanosomes invade the CNS
- Author
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Christian Boßelmann, Hartwig Wolburg, Michael Duszenko, Andreas Meiwes, and Stefan Mogk
- Subjects
Trypanosoma ,Central nervous system ,Brain ,Disease ,Biology ,Infectious Diseases ,medicine.anatomical_structure ,Blood ,Meninges ,Trypanosomiasis, African ,Drug development ,Invasion process ,Blood-Brain Barrier ,Immunology ,Chronic Disease ,medicine ,Animals ,Humans ,Parasitology ,Blood csf barrier ,Cerebrospinal Fluid - Abstract
African trypanosomes induce sleeping sickness. Although it is clear that this parasite moves from the blood to the central nervous system (CNS) to induce the second stage of the disease, little is known about the molecular details of this process. Considering new findings of the trypanosome localization, this opinion paper will summarize the current knowledge about CNS infection, propose a different perception of the invasion process, and discuss possible consequences for drug development.
- Published
- 2014
17. Serial crystallography on in vivo grown microcrystals using synchrotron radiation
- Author
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Lars Redecke, B.P. Sommer, Oleksandr Yefanov, Thomas R. Schneider, Gleb Bourenkov, Henry N. Chapman, Dominik Oberthür, Stefan Mogk, Francesco Stellato, Cornelius Gati, Christian Betzel, M. Klinge, Dirk Rehders, and Michael Duszenko
- Subjects
Diffraction ,Crystallography ,Materials science ,in vivo grown microcrystals ,Settore FIS/07 ,Resolution (electron density) ,Synchrotron radiation ,General Chemistry ,Crystal structure ,Condensed Matter Physics ,Laser ,Research Papers ,Biochemistry ,Synchrotron ,law.invention ,Crystal ,QD901-999 ,law ,protein microcrystallography ,Femtosecond ,serial crystallography ,General Materials Science ,ddc:530 - Abstract
The structure solution of T. brucei cathepsin B from 80 in vivo grown crystals with an average volume of 9 µm3 obtained by serial synchrotron crystallography at a microfocus beamline is reported., Crystal structure determinations of biological macromolecules are limited by the availability of sufficiently sized crystals and by the fact that crystal quality deteriorates during data collection owing to radiation damage. Exploiting a micrometre-sized X-ray beam, high-precision diffractometry and shutterless data acquisition with a pixel-array detector, a strategy for collecting data from many micrometre-sized crystals presented to an X-ray beam in a vitrified suspension is demonstrated. By combining diffraction data from 80 Trypanosoma brucei procathepsin B crystals with an average volume of 9 µm3, a complete data set to 3.0 Å resolution has been assembled. The data allowed the refinement of a structural model that is consistent with that previously obtained using free-electron laser radiation, providing mutual validation. Further improvements of the serial synchrotron crystallography technique and its combination with serial femtosecond crystallography are discussed that may allow the determination of high-resolution structures of micrometre-sized crystals.
- Published
- 2014
18. Natively inhibited Trypanosoma brucei cathepsin B structure determined by using an X-ray laser
- Author
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R. Bruce Doak, Matthias Frank, Francesco Stellato, Gergely Katona, Richard Neutze, David Arnlund, Marc Messerschmidt, Dingjie Wang, M. Marvin Seibert, Dirk Rehders, Richard A. Kirian, Robert L. Shoeman, Thomas R. M. Barends, Henry N. Chapman, Nicusor Timneanu, Andrew Aquila, Ingo Grotjohann, Tzu-Chiao Chao, Nadia A. Zatsepin, Torsten Barth, Petra Fromme, Linda C. Johansson, Saša Bajt, Michael Duszenko, Christian Betzel, Mark S. Hunter, Sébastien Boutet, Lars Redecke, Chris Kupitz, Karol Nass, Andrew V. Martin, Daniel P. DePonte, Lorenzo Galli, Stefan Mogk, Garth J. Williams, Lukas Lomb, Rudolf Koopmann, Mengning Liang, Holger Fleckenstein, Uwe Weierstall, Anton Barty, Ilme Schlichting, Michael J. Bogan, Stephan Kassemeyer, Jan Steinbrener, Raimund Fromme, Carl Caleman, John C. H. Spence, and Thomas A. White
- Subjects
Models, Molecular ,Glycosylation ,Protein Conformation ,Molecular Sequence Data ,Trypanosoma brucei brucei ,Protozoan Proteins ,Biology ,Trypanosoma brucei ,Spodoptera ,Crystallography, X-Ray ,Cathepsin B ,In vivo ,Catalytic Domain ,Sf9 Cells ,Animals ,Angstrom ,Amino Acid Sequence ,Host protein ,Enzyme Precursors ,Multidisciplinary ,X-Rays ,biology.organism_classification ,Protozoan parasite ,Cysteine protease ,Biochemistry ,Fatal disease ,Crystallization - Abstract
Diffraction Before Destruction A bottleneck in x-ray crystallography is the growth of well-ordered crystals large enough to obtain high-resolution diffraction data within an exposure that limits radiation damage. Serial femtosecond crystallography promises to overcome these constraints by using short intense pulses that out-run radiation damage. A stream of crystals is flowed across the free-electron beam and for each pulse, diffraction data is recorded from a single crystal before it is destroyed. Redecke et al. (p. 227 , published online 29 November; see the Perspective by Helliwell ) used this technique to determine the structure of an enzyme from Trypanosoma brucei , the parasite that causes sleeping sickness, from micron-sized crystals grown within insect cells. The structure shows how this enzyme, which is involved in degradation of host proteins, is natively inhibited prior to activation, which could help in the development of parasite-specific inhibitors.
- Published
- 2013
19. Drug development against sleeping sickness: old wine in new bottles?
- Author
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Stefan Mogk, Michael Duszenko, A. Gray, M. Huber, M. Scholze, Celestin Nzanzu Mudogo, B.P. Sommer, Andreas Meiwes, and Jasmin Stein
- Subjects
Drug ,Trypanosoma ,media_common.quotation_subject ,Druggability ,Disease ,Pharmacology ,Biology ,Bioinformatics ,Biochemistry ,Cell density ,Drug Discovery ,medicine ,Animals ,Humans ,African trypanosomiasis ,media_common ,Life Cycle Stages ,Drug discovery ,Organic Chemistry ,medicine.disease ,Trypanocidal Agents ,Trypanosomiasis, African ,Drug development ,Molecular Medicine ,Energy Metabolism ,Trypanosomiasis - Abstract
Atoxyl, the first medicinal drug against human African trypanosomiasis (HAT), also known as sleeping sickness, was applied more than 100 years ago. Ever since, the search for more effective, more specific and less toxic drugs continued, leading to a set of compounds currently in use against this devastating disease. Unfortunately, none of these medicines fulfill modern pharmaceutical requirements and may be considered as therapeutic ultima ratio due to the many, often severe side effects. Starting with a historic overview on drug development against HAT, we present a selection of trypanosome specific pathways and enzymes considered as highly potent druggable targets. In addition, we describe cellular mechanisms the parasite uses for differentiation and cell density regulation and present our considerations how interference with these steps, elementary for life cycle progression and infection, may lead to new aspects of drug development. Finally we refer to our recent work about CNS infection that offers novel insights in how trypanosomes hide in an immune privileged area to establish a chronic state of the disease, thereby considering new ways for drug application. Depressingly, HAT specific drug development has failed over the last 30 years to produce better suited medicine. However, unraveling of parasite-specific pathways and cellular behavior together with the ability to produce high resolution structures of essential parasite proteins by X-ray crystallography, leads us to the optimistic view that development of an ultimate drug to eradicate sleeping sickness from the globe might just be around the corner.
- Published
- 2012
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