82 results on '"Jiří Brynda"'
Search Results
2. X-ray structure of human aldo–keto reductase 1C3 in complex with a bile acid fused tetrazole inhibitor: experimental validation, molecular docking and structural analysis
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Maja A. Marinović, Sofija S. Bekić, Michael Kugler, Jiří Brynda, Jana Škerlová, Dušan Đ. Škorić, Pavlína Řezáčová, Edward T. Petri, and Andjelka S. Ćelić
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Pharmacology ,Organic Chemistry ,Drug Discovery ,Pharmaceutical Science ,Molecular Medicine ,Biochemistry - Abstract
Steroid bile acid fused tetrazoles were screened for ability to inhibit aldo–keto reductase 1C3 (AKR1C3), a target for cancer treatment. The X-ray structure of AKR1C3 in complex with a bile acid tetrazole inhibitor is presented.
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- 2023
3. Design, Synthesis, and Biochemical and Biological Evaluation of Novel 7-Deazapurine Cyclic Dinucleotide Analogues as STING Receptor Agonists
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Zdeněk Vavřina, Pavla Perlíková, Nemanja Milisavljević, Florian Chevrier, Miroslav Smola, Joshua Smith, Milan Dejmek, Vojtěch Havlíček, Miloš Buděšínský, Radek Liboska, Lenka Vaneková, Jiří Brynda, Evzen Boura, Pavlína Řezáčová, Michal Hocek, and Gabriel Birkuš
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Mice ,Drug Discovery ,Animals ,Humans ,Membrane Proteins ,Cytokines ,Molecular Medicine ,Interferons ,Nucleotides, Cyclic ,Ligands ,Nucleotidyltransferases - Abstract
Cyclic dinucleotides (CDNs) are second messengers that activate stimulator of interferon genes (STING). The cGAS-STING pathway plays a promising role in cancer immunotherapy. Here, we describe the synthesis of CDNs containing 7-substituted 7-deazapurine moiety. We used mouse cyclic GMP-AMP synthase and bacterial dinucleotide synthases for the enzymatic synthesis of CDNs. Alternatively, 7-(het)aryl 7-deazapurine CDNs were prepared by Suzuki-Miyaura cross-couplings. New CDNs were tested in biochemical and cell-based assays for their affinity to human STING. Eight CDNs showed better activity than 2'3'
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- 2022
4. Mercury removal in coal-fired power plants, possibilities how to attain very low emissions and minimization of hazardous waste stream
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Karel Svoboda, Michael Pohořelý, Tomáš Ružovič, Václav Veselý, Jiří Brynda, Boleslav Zach, and Michal Šyc
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Fuel Technology ,Environmental Engineering ,Renewable Energy, Sustainability and the Environment ,Energy Engineering and Power Technology ,Geotechnical Engineering and Engineering Geology - Abstract
Emissions of toxic heavy metals (HMs), as Hg, As, Cd, Pb, etc., and some harmful compounds of F, Se, and B are related to waste streams from coal-fired power plants (CFPP). Coal/lignite combustion, due to relatively high content of ash, sulfur, and chlorine, generates in flue gas cleaning processes tremendous amount of fly ash, CaSO4 and CaCl2. Measures for minimization of Hg- and NOx-emissions (e.g. addition of bromides and NH3) change properties of fly ash, wastewater and speciation/partition of HMs. Wet flue gas desulfurization (FGD) consumes high amount of fresh water and generates harmful wastewater with water soluble salts. The planned, more stringent limits on emissions of dust, Hg, HCl, HF, SO2, etc. in CFPP will increase contents of polluting compounds in solid and liquid waste streams. We critically assess possibilities, measures and obstacles for higher efficiency of Hg and HMs removal from flue gas in CFPP, together with efficient removal of other pollutants including mutual influences and interrelations. The fates of mercury, selected harmful HMs, and some other pollutants in waste streams from wet FGD are critically analyzed and discussed. Non-toxic, stable forms of mercury (e.g. HgS) and other HMs in solid waste should be preferred. Schemes and measures for minimization of emissions and hazardous waste streams from air pollution control (APC) are compared and discussed for three selected technologies of coal combustion with different methods of gas cleaning.
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- 2022
5. B-H⋯π and C-H⋯π interactions in protein-ligand complexes: carbonic anhydrase II inhibition by carborane sulfonamides
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Jindřich Fanfrlík, Jiří Brynda, Michael Kugler, Martin Lepšík, Klára Pospíšilová, Josef Holub, Drahomír Hnyk, Jan Nekvinda, Bohumír Grüner, and Pavlína Řezáčová
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General Physics and Astronomy ,Physical and Theoretical Chemistry - Abstract
Among non-covalent interactions, B-H⋯π and C-H⋯π hydrogen bonding is rather weak and less studied. Nevertheless, since both can affect the energetics of protein-ligand binding, their understanding is an important prerequisite for reliable predictions of affinities. Through a combination of high-resolution X-ray crystallography and quantum-chemical calculations on carbonic anhydrase II/carborane-based inhibitor systems, this paper provides the first example of B-H⋯π hydrogen bonding in a protein-ligand complex. It shows that the B-H⋯π interaction is stabilized by dispersion, followed by electrostatics. Furthermore, it demonstrates that the similar C-H⋯π interaction is twice as strong, with a slightly smaller contribution of dispersion and a slightly higher contribution of electrostatics. Such a detailed insight will facilitate the rational design of future protein ligands, controlling these types of non-covalent interactions.
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- 2023
6. Structural insight into DNA recognition by bacterial transcriptional regulators of the SorC/DeoR family
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Markéta Šoltysová, Jana Škerlová, Pavlína Řezáčová, Milan Fábry, Jiří Brynda, and Irena Sieglová
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Models, Molecular ,Binding Sites ,biology ,Protein Conformation ,Regulator ,DNA ,Bacillus subtilis ,Computational biology ,Crystallography, X-Ray ,biology.organism_classification ,DNA-Binding Proteins ,chemistry.chemical_compound ,Quorum sensing ,Bacterial Proteins ,chemistry ,Structural Biology ,Bacterial transcription ,Gene ,Dna recognition ,Protein Binding - Abstract
The SorC/DeoR family is a large family of bacterial transcription regulators that are involved in the control of carbohydrate metabolism and quorum sensing. To understand the structural basis of DNA recognition, structural studies of two functionally characterized SorC/DeoR family members from Bacillus subtilis were performed: the deoxyribonucleoside regulator bsDeoR and the central glycolytic genes regulator bsCggR. Each selected protein represents one of the subgroups that are recognized within the family. Crystal structures were determined of the N-terminal DNA-binding domains of bsDeoR and bsCggR in complex with DNA duplexes representing the minimal operator sequence at resolutions of 2.3 and 2.1 Å, respectively. While bsDeoRDBD contains a homeodomain-like HTH-type domain, bsCggRDBD contains a winged helix–turn–helix-type motif. Both proteins form C2-symmetric dimers that recognize two consecutive major grooves, and the protein–DNA interactions have been analyzed in detail. The crystal structures were used to model the interactions of the proteins with the full DNA operators, and a common mode of DNA recognition is proposed that is most likely to be shared by other members of the SorC/DeoR family.
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- 2021
7. Re-emerging Aspartic Protease Targets: Examining Cryptococcus neoformans Major Aspartyl Peptidase 1 as a Target for Antifungal Drug Discovery
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Marcin Drąg, Jiří Brynda, Robin Kryštůfek, Jan Konvalinka, Eva Tloušt’ová, Martin Hradilek, Michael J. Boucher, Pavel Šácha, Justyna Grzymska, Pavlína Řezáčová, Miroslav Hájek, Jana Starková, Wioletta Rut, Charles S. Craik, Hiten D. Madhani, and Pavel Majer
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Proteases ,Antifungal Agents ,Aspartic Acid Proteases ,Medicinal & Biomolecular Chemistry ,medicine.medical_treatment ,Drug Evaluation, Preclinical ,Antifungal drug ,Molecular Dynamics Simulation ,Crystallography, X-Ray ,Article ,Substrate Specificity ,Microbiology ,Fungal Proteins ,Structure-Activity Relationship ,Medicinal and Biomolecular Chemistry ,HIV Protease ,Catalytic Domain ,Drug Discovery ,medicine ,HIV Protease Inhibitor ,Protease inhibitor (pharmacology) ,Cryptococcus neoformans ,Binding Sites ,Crystallography ,Protease ,biology ,Chemistry ,Organic Chemistry ,Fungi ,HIV ,Pharmacology and Pharmaceutical Sciences ,medicine.disease ,biology.organism_classification ,Aspartyl Peptidase ,Preclinical ,Recombinant Proteins ,Infectious Diseases ,5.1 Pharmaceuticals ,6.1 Pharmaceuticals ,Cryptococcosis ,X-Ray ,HIV/AIDS ,Drug Evaluation ,Molecular Medicine ,Infection - Abstract
Cryptococcosis is an invasive infection that accounts for 15% of AIDS-related fatalities. Still, treating cryptococcosis remains a significant challenge due to the poor availability of effective antifungal therapies and emergence of drug resistance. Interestingly, protease inhibitor components of antiretroviral therapy regimens have shown some clinical benefits in these opportunistic infections. We investigated Major aspartyl peptidase 1 (May1), a secreted Cryptococcus neoformans protease, as a possible target for the development of drugs that act against both fungal and retroviral aspartyl proteases. Here, we describe the biochemical characterization of May1, present its high-resolution X-ray structure, and provide its substrate specificity analysis. Through combinatorial screening of 11,520 compounds, we identified a potent inhibitor of May1 and HIV protease. This dual-specificity inhibitor exhibits antifungal activity in yeast culture, low cytotoxicity, and low off-target activity against host proteases and could thus serve as a lead compound for further development of May1 and HIV protease inhibitors.
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- 2021
8. The Influence of SO2 and HCl Concentrations on the Consumption of Sodium Bicarbonate during Flue Gas Treatment
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Boleslav Zach, Radovan Šomplák, M. Punčochář, Jiří Brynda, Jaroslav Moško, Michael Pohořelý, Karel Svoboda, and Michal Šyc
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Flue gas ,Treatment system ,Sodium bicarbonate ,Sorbent ,General Chemical Engineering ,Inorganic chemistry ,Energy Engineering and Power Technology ,02 engineering and technology ,021001 nanoscience & nanotechnology ,Target concentration ,chemistry.chemical_compound ,Fuel Technology ,020401 chemical engineering ,chemistry ,0204 chemical engineering ,0210 nano-technology ,Sodium carbonate - Abstract
The paper focuses on the competitive influence of SO₂ and HCl during their removal from flue gas by sodium bicarbonate. The experimental investigation was conducted at a unique combination of conditions: the experiments were conducted on a large scale, with real flue gas, and the removal of SO₂ and HCl was studied simultaneously. The aim was to ensure that experimental conditions are reasonably realistic and therefore transferable to practice. The results from experiments at various temperatures within the range from 170 to 255 °C did not indicate a significant influence of temperature. The sorbent conversion varied significantly with the lowest achieved conversion of 26% and the highest conversion of 85%. There was a clear trend showing that higher concentrations of HCl make it more difficult to achieve the selected target concentration of SO₂ in clean flue gas, suggesting the preferred reaction of formed sodium carbonate with HCl. The influence of HCl on the removal of SO₂ was quantified and is presented in the form of regression models. The found regressors were the initial concentrations of HCl and SO₂ in raw flue gas and the target concentration of SO₂ in the clean flue gas. The regression models are in good agreement with the experimental data and can be used for the prediction of the behavior of a flue gas treatment system or its optimization.
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- 2021
9. Directionality of light absorption and emission in representative fluorescent proteins
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Jitka Myskova, Alexey Bondar, Jiří Brynda, Josef Lazar, Olga Rybakova, and Petro Khoroshyy
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Physics ,Dipole ,Bioelectronics ,Multidisciplinary ,Optics ,business.industry ,Transition dipole moment ,Directionality ,Polarization Microscopy ,mCherry ,business ,Fluorescence ,Excitation - Abstract
Fluorescent molecules are like antennas: The rate at which they absorb light depends on their orientation with respect to the incoming light wave, and the apparent intensity of their emission depends on their orientation with respect to the observer. However, the directions along which the most important fluorescent molecules in biology, fluorescent proteins (FPs), absorb and emit light are generally not known. Our optical and X-ray investigations of FP crystals have now allowed us to determine the molecular orientations of the excitation and emission transition dipole moments in the FPs mTurquoise2, eGFP, and mCherry, and the photoconvertible FP mEos4b. Our results will allow using FP directionality in studies of molecular and biological processes, but also in development of novel bioengineering and bioelectronics applications.
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- 2020
10. Direct Introduction of an Alkylsulfonamido Group on C‐sites of Isomeric Dicarba‐ closo ‐dodecaboranes: The Influence of Stereochemistry on Inhibitory Activity against the Cancer‐Associated Carbonic Anhydrase IX Isoenzyme
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K. Pospisilova, Pavlína Řezáčová, Jan Nekvinda, Zdeňka Růžičková, Bohumír Grüner, Suzan El Anwar, Jiří Brynda, Josef Holub, and M. Kugler
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Carbonic Anhydrase I ,Stereochemistry ,Crystal structure ,010402 general chemistry ,Inhibitory postsynaptic potential ,01 natural sciences ,Isozyme ,Catalysis ,Structure-Activity Relationship ,Antigens, Neoplasm ,Neoplasms ,Carbonic anhydrase ,Humans ,Carbonic Anhydrase IX ,Carbonic Anhydrase Inhibitors ,chemistry.chemical_classification ,biology ,010405 organic chemistry ,Organic Chemistry ,General Chemistry ,In vitro ,0104 chemical sciences ,Sulfonamide ,Isoenzymes ,Enzyme ,chemistry ,biology.protein ,Selectivity - Abstract
Carbonic anhydrase IX (CA IX), a tumor-associated metalloenzyme, represents a validated target for cancer therapy and diagnostics. Herein, we report the inhibition properties of isomeric families of sulfonamidopropyl-dicarba-closo-dodecaboranes group(s) prepared using a new direct five-step synthesis from the corresponding parent cages. The protocol offers a reliable solution for synthesis of singly and doubly substituted dicarba-closo-dodecaboranes with a different geometric position of carbon atoms. The closo-compounds from the ortho- and meta-series were then degraded to corresponding 11-vertex dicarba-nido-undecaborate(1-) anions. All compounds show in vitro enzymatic activity against CA IX in the low nanomolar or subnanomolar range. This is accompanied by clear isomer dependence of the inhibition constant (Ki ) and selectivity towards CA IX. Decreasing trends in Ki and selectivity index (SI ) values are observed with increasing separation of the cage carbon atoms. Interactions of compounds with the active sites of CA IX were explored with X-ray crystallography, and eight high-resolution crystal structures uncovered the structural basis of inhibition potency and selectivity.
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- 2020
11. Identification of Novel Carbonic Anhydrase IX Inhibitors Using High-Throughput Screening of Pooled Compound Libraries by DNA-Linked Inhibitor Antibody Assay (DIANA)
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Jiří Brynda, Ullrich Jahn, Milan Potáček, Anna Hlavačková, Vlastimil Král, M. Kugler, Pavlína Řezáčová, Jiří Schimer, Milan Dejmek, Pavel Šácha, Jan Tykvart, Jan Konvalinka, Lukáš Tenora, Václav Navrátil, Jitka Zemanova, and Pavel Majer
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High-throughput screening ,01 natural sciences ,Biochemistry ,Analytical Chemistry ,03 medical and health sciences ,chemistry.chemical_compound ,Antigens, Neoplasm ,Catalytic Domain ,Humans ,Potency ,Carbonic Anhydrase IX ,Carbonic Anhydrase Inhibitors ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,biology ,010405 organic chemistry ,Chemistry ,Drug discovery ,DNA ,High-Throughput Screening Assays ,3. Good health ,0104 chemical sciences ,Molecular Docking Simulation ,Enzyme ,Pharmaceutical Preparations ,Enzyme inhibitor ,biology.protein ,Molecular Medicine ,Biological Assay ,Antibody ,Biotechnology - Abstract
The DNA-linked inhibitor antibody assay (DIANA) has been recently validated for ultrasensitive enzyme detection and for quantitative evaluation of enzyme inhibitor potency. Here we present its adaptation for high-throughput screening of human carbonic anhydrase IX (CAIX), a promising drug and diagnostic target. We tested DIANA's performance by screening a unique compound collection of 2816 compounds consisting of lead-like small molecules synthesized at the Institute of Organic Chemistry and Biochemistry (IOCB) Prague ("IOCB library"). Additionally, to test the robustness of the assay and its potential for upscaling, we screened a pooled version of the IOCB library. The results from the pooled screening were in agreement with the initial nonpooled screen with no lost hits and no false positives, which shows DIANA's potential to screen more than 100,000 compounds per day.All DIANA screens showed a high signal-to-noise ratio with a Z' factor of0.89. The DIANA screen identified 13 compounds with
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- 2020
12. Highly potent inhibitors of cathepsin K with a differently positioned cyanohydrazide warhead: structural analysis of binding mode to mature and zymogen-like enzymes
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Jakub Benýšek, Michal Buša, Petra Rubešová, Jindřich Fanfrlík, Martin Lepšík, Jiří Brynda, Zuzana Matoušková, Ulrike Bartz, Martin Horn, Michael Gütschow, and Michael Mareš
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Models, Molecular ,Pharmacology ,Dose-Response Relationship, Drug ,Molecular Structure ,Cathepsin K ,General Medicine ,Structure-Activity Relationship ,Hydrazines ,Nitriles ,ddc:540 ,Drug Discovery ,Tumor Cells, Cultured ,Humans ,Protease Inhibitors - Abstract
Cathepsin K (CatK) is a target for the treatment of osteoporosis, arthritis, and bone metastasis. Peptidomimetics with a cyanohydrazide warhead represent a new class of highly potent CatK inhibitors; however, their binding mechanism is unknown. We investigated two model cyanohydrazide inhibitors with differently positioned warheads: an azadipeptide nitrile G��1303 and a 3-cyano-3-aza-��-amino acid G��2602. Crystal structures of their covalent complexes were determined with mature CatK as well as a zymogen-like activation intermediate of CatK. Binding mode analysis, together with quantum chemical calculations, revealed that the extraordinary picomolar potency of G��2602 is entropically favoured by its conformational flexibility at the nonprimed-primed subsites boundary. Furthermore, we demonstrated by live cell imaging that cyanohydrazides effectively target mature CatK in osteosarcoma cells. Cyanohydrazides also suppressed the maturation of CatK by inhibiting the autoactivation of the CatK zymogen. Our results provide structural insights for the rational design of cyanohydrazide inhibitors of CatK as potential drugs.
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- 2022
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13. Thermodynamic and structural characterization of an optimized peptide-based inhibitor of the influenza polymerase PA-PB1 subunit interaction
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Kateřina Radilová, Václav Zima, Michal Kráľ, Aleš Machara, Pavel Majer, Jan Hodek, Jan Weber, Jiří Brynda, Timotej Strmeň, Jan Konvalinka, and Milan Kožíšek
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Pharmacology ,Virology ,Influenza, Human ,Humans ,Thermodynamics ,RNA-Dependent RNA Polymerase ,Peptides ,Protein Binding - Abstract
Influenza virus causes severe respiratory infection in humans. Current antivirotics target three key proteins in the viral life cycle: neuraminidase, the M2 channel and the endonuclease domain of RNA-dependent-RNA polymerase. Due to the development of novel pandemic strains, additional antiviral drugs targetting different viral proteins are still needed. The protein-protein interaction between polymerase subunits PA and PB1 is one such possible target. We recently identified a modified decapeptide derived from the N-terminus of the PB1 subunit with high affinity for the C-terminal part of the PA subunit. Here, we optimized its amino acid hotspots to maintain the inhibitory potency and greatly increase peptide solubility. This allowed thermodynamic characterization of peptide binding to PA. Solving the X-ray structure of the peptide-PA complex provided structural insights into the interaction. Additionally, we optimized intracellular delivery of the peptide using a bicyclic strategy that led to improved inhibition in cell-based assays.
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- 2022
14. Metallacarborane Sulfamides: Unconventional, Specific, and Highly Selective Inhibitors of Carbonic Anhydrase IX
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Soňa Gurská, Bohumír Grüner, Jana Štěpánková, Marian Hajduch, K. Pospisilova, Alice Nova, Pavlína Řezáčová, Václav Šícha, Milan Fábry, Vlastimil Mašek, Petr Džubák, Stanislava Matějková, Jan Nekvinda, Petr Pachl, Josef Holub, Viswanath Das, Jiří Brynda, Barbora Lišková, Martina Medvedíková, Vlastimil Král, and M. Kugler
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Models, Molecular ,Gene isoform ,Mice ,Catalytic Domain ,Cell Line, Tumor ,Drug Discovery ,medicine ,Animals ,Humans ,Doxorubicin ,Boranes ,Carbonic Anhydrase IX ,Carbonic Anhydrase Inhibitors ,chemistry.chemical_classification ,biology ,Chemistry ,Active site ,Biological Transport ,Drug Synergism ,Amides ,Xenograft Model Antitumor Assays ,Transmembrane protein ,Cytosol ,Enzyme ,Cell culture ,Drug Design ,biology.protein ,Cancer research ,Molecular Medicine ,Syngenic ,medicine.drug - Abstract
Carbonic anhydrase IX (CAIX) is a transmembrane enzyme that regulates pH in hypoxic tumors and promotes tumor cell survival. Its expression is associated with the occurrence of metastases and poor prognosis. Here, we present nine derivatives of the cobalt bis(dicarbollide)(1-) anion substituted at the boron or carbon sites by alkysulfamide group(s) as highly specific and selective inhibitors of CAIX. Interactions of these compounds with the active site of CAIX were explored on the atomic level using protein crystallography. Two selected derivatives display subnanomolar or picomolar inhibition constants and high selectivity for the tumor-specific CAIX over cytosolic isoform CAII. Both derivatives had a time-dependent effect on the growth of multicellular spheroids of HT-29 and HCT116 colorectal cancer cells, facilitated penetration and/or accumulation of doxorubicin into spheroids, and displayed low toxicity and showed promising pharmacokinetics and a significant inhibitory effect on tumor growth in syngenic breast 4T1 and colorectal HT-29 cancer xenotransplants.
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- 2019
15. Capturing a dynamically interacting inhibitor by paramagnetic NMR spectroscopy
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Klára Grantz-Šašková, Ladislav Benda, Jan Konvalinka, Pavel Srb, Václav Šícha, Jiří Brynda, Michal Svoboda, Bohumír Grüner, Martin Lepšík, Ján Tarábek, Vaclav Veverka, and Pavlína Řezáčová
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Magnetic Resonance Spectroscopy ,Materials science ,Protein Conformation ,General Physics and Astronomy ,02 engineering and technology ,Molecular Dynamics Simulation ,Ligands ,010402 general chemistry ,01 natural sciences ,Paramagnetism ,Molecular dynamics ,Protein structure ,HIV Protease ,Physical and Theoretical Chemistry ,Boranes ,Quantitative Biology::Biomolecules ,Intermolecular force ,Nuclear magnetic resonance spectroscopy ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Characterization (materials science) ,NMR spectra database ,Metals ,Chemical physics ,Quantum Theory ,Diamagnetism ,0210 nano-technology ,Protein Binding - Abstract
Transient and fuzzy intermolecular interactions are fundamental to many biological processes. Despite their importance, they are notoriously challenging to characterize. Effects induced by paramagnetic ligands in the NMR spectra of interacting biomolecules provide an opportunity to amplify subtle manifestations of weak intermolecular interactions observed for diamagnetic ligands. Here, we present an approach to characterizing dynamic interactions between a partially flexible dimeric protein, HIV-1 protease, and a metallacarborane-based ligand, a system for which data obtained by standard NMR approaches do not enable detailed structural interpretation. We show that for the case where the experimental data are significantly averaged to values close to zero the standard fitting of pseudocontact shifts cannot provide reliable structural information. We based our approach on generating a large ensemble of full atomic models, for which the experimental data can be predicted, ensemble averaged and finally compared to the experiment. We demonstrate that a combination of paramagnetic NMR experiments, quantum chemical calculations, and molecular dynamics simulations offers a route towards structural characterization of dynamic protein-ligand complexes.
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- 2019
16. Inhibitors of CA IX Enzyme Based on Polyhedral Boron Compounds
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Marian Hajduch, Bohumír Grüner, Jan Nekvinda, K. Pospisilova, Pavlína Řezáčová, Václav Šícha, Milan Fábry, Josef Holub, Viswanath Das, Jiří Brynda, Vlastimil Král, M. Kugler, Suzan El Anwar, and Václav Kašička
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Boron Compounds ,Combination therapy ,chemistry.chemical_element ,Molecular Dynamics Simulation ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Structure-Activity Relationship ,Carbonic anhydrase ,Neoplasms ,Organometallic Compounds ,Humans ,Boron ,Structural motif ,Carbonic Anhydrase IX ,Carbonic Anhydrase Inhibitors ,Molecular Biology ,chemistry.chemical_classification ,Sulfonamides ,Binding Sites ,biology ,010405 organic chemistry ,Organic Chemistry ,Active site ,Combinatorial chemistry ,0104 chemical sciences ,Enzyme ,chemistry ,Drug development ,biology.protein ,Molecular Medicine ,Carborane - Abstract
This review describes recent progress in the design and development of inhibitors of human carbonic anhydrase IX (CA IX) based on space-filling carborane and cobalt bis(dicarbollide) clusters. CA IX enzyme is known to play a crucial role in cancer cell proliferation and metastases. The new class of potent and selective CA IX inhibitors combines the structural motif of a bulky inorganic cluster with an alkylsulfamido or alkylsulfonamido anchor group for Zn2+ ion in the enzyme active site. Detailed structure-activity relationship (SAR) studies of a large series containing 50 compounds uncovered structural features of the cluster-containing inhibitors that are important for efficient and selective inhibition of CA IX activity. Preclinical evaluation of selected compounds revealed low toxicity, favorable pharmacokinetics and ability to reduce tumor growth. Cluster-containing inhibitors of CA IX can thus be considered as promising candidates for drug development and/or for combination therapy in boron neutron capture therapy (BNCT).
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- 2021
17. Structural determinants for subnanomolar inhibition of the secreted aspartic protease Sapp1p from
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Jiří, Dostál, Jiří, Brynda, Lucie, Vaňková, Syeda Rehana, Zia, Iva, Pichová, Olga, Heidingsfeld, and Martin, Lepšík
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Models, Molecular ,crystal structure ,Candida parapsilosis ,Inhibitor ,Dose-Response Relationship, Drug ,Molecular Structure ,Fungal Proteins ,Structure-Activity Relationship ,noncovalent interactions ,peptidomimetics ,hydrogen bonds ,Aspartic Acid Endopeptidases ,Protease Inhibitors ,Research Article ,Research Paper - Abstract
Pathogenic Candida albicans yeasts frequently cause infections in hospitals. Antifungal drugs lose effectiveness due to other Candida species and resistance. New medications are thus required. Secreted aspartic protease of C. parapsilosis (Sapp1p) is a promising target. We have thus solved the crystal structures of Sapp1p complexed to four peptidomimetic inhibitors. Three potent inhibitors (Ki: 0.1, 0.4, 6.6 nM) resembled pepstatin A (Ki: 0.3 nM), a general aspartic protease inhibitor, in terms of their interactions with Sapp1p. However, the weaker inhibitor (Ki: 14.6 nM) formed fewer nonpolar contacts with Sapp1p, similarly to the smaller HIV protease inhibitor ritonavir (Ki: 1.9 µM), which, moreover, formed fewer H-bonds. The analyses have revealed the structural determinants of the subnanomolar inhibition of C. parapsilosis aspartic protease. Because of the high similarity between Saps from different Candida species, these results can further be used for the design of potent and specific Sap inhibitor-based antimycotic drugs.
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- 2021
18. Structure-activity relationship and biochemical evaluation of novel fibroblast activation protein and prolyl endopeptidase inhibitors with α-ketoamide warheads
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Tereza Ormsby, Pavel Majer, Natan Sidej, Jiří Brynda, Lenka Poštová Slavětínská, Adéla Šimková, Jana Beranová, Pavel Šácha, and Jan Konvalinka
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Pharmacology ,chemistry.chemical_classification ,Molecular Structure ,Peptidomimetic ,Chemistry ,Organic Chemistry ,Peptide ,General Medicine ,Fibroblasts ,Mice ,Structure-Activity Relationship ,Prolyl endopeptidase ,Fibroblast activation protein, alpha ,Biochemistry ,In vivo ,Drug Discovery ,medicine ,Potency ,Structure–activity relationship ,Animals ,Humans ,Cytotoxicity ,Prolyl Oligopeptidases ,medicine.drug - Abstract
Peptidomimetic inhibitors of fibroblast activation protein (FAP) are regarded as promising tools for tumor targeting in vivo. Even though several peptidomimetic compounds with nanomolar potency have been described, broad chemical space for further modification remained unexplored. Therefore, we set to analyze the structure-activity relationship (SAR) of pseudopeptide compound series with α-ketoamide warheads in order to explore the contributions of the P1′ and P2′ moieties to the inhibitory potency. A series of novel inhibitors bearing varied P1′ and/or P2’ moieties was synthesized by combining a Passerini reaction-Amine Deprotection-Acyl Migration (PADAM) approach with peptide coupling and subsequent oxidation. The resulting compounds inhibited FAP and the related prolyl endopeptidase (PREP) with potencies in the nanomolar to sub-nanomolar range. The most potent FAP inhibitor IOCB22-AP446 (6d, IC50 = 89 pM) had about 36 -fold higher inhibition potency than the most potent inhibitor published to date. The compounds were selective over FAP's closest homolog DPP-IV, were stable in human and mouse plasma and in mouse microsomes, and displayed minimal cytotoxicity in tissue cultures.
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- 2021
19. Determination of acidity constants, ionic mobilities, and hydrodynamic radii of carborane-based inhibitors of carbonic anhydrases by capillary electrophoresis
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Bohumír Grűner, Jiří Brynda, Veronika Šolínová, Josef Holub, Václav Šícha, and Václav Kašička
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Clinical Biochemistry ,Ionic bonding ,02 engineering and technology ,01 natural sciences ,Biochemistry ,Acid dissociation constant ,Analytical Chemistry ,Divalent ,Ion ,Capillary electrophoresis ,Carbonic Anhydrases ,chemistry.chemical_classification ,Ions ,010401 analytical chemistry ,Electrophoresis, Capillary ,Hydrogen-Ion Concentration ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Electrophoresis ,chemistry ,Ionic strength ,Hydrodynamics ,Carborane ,Physical chemistry ,0210 nano-technology ,Acids - Abstract
Capillary electrophoresis (CE) has been applied for determination of the thermodynamic acidity constants (pKa ) of the sulfamidoalkyl and sulfonamidoalkyl groups, the actual and limiting ionic mobilities and hydrodynamic radii of important compounds, eight carborane-based inhibitors of carbonic anhydrases, which are potential new anticancer drugs. Two types of carboranes were investigated, (i) icosahedral cobalt bis(dicarbollide)(1-) ion with sulfamidoalkyl moieties, and (ii) 7,8-nido-dicarbaundecaborate with sulfonamidoalkyl side chains. First, the mixed acidity constants, pKamix , of the sulfamidoalkyl and sulfonamidoalkyl groups of the above carboranes and their actual ionic mobilities were determined by nonlinear regression analysis of the pH dependences of their effective electrophoretic mobility measured by capillary electrophoresis in the pH range 8.00-12.25, at constant ionic strength (25 mM), and constant temperature (25°C). Second, the pKamix were recalculated to the thermodynamic pKa s using the Debye-Huckel theory. The sulfamidoalkyl and sulfonamidoalkyl groups were found to be very weakly acidic with the pKa s in the range 10.78-11.45 depending on the type of carborane cluster and on the position and length of the alkyl chain on the carborane scaffold. These pKa s were in a good agreement with the pKa s (10.67-11.27) obtained by new program AnglerFish (freeware at https://echmet.natur.cuni.cz), which provides thermodynamic pKa s and limiting ionic mobilities directly from the raw CE data. The absolute values of the limiting ionic mobilities of univalent and divalent carborane anions were in the range 18.3-27.8 TU (Tiselius unit, 1 × 10-9 m2 /Vs), and 36.4-45.9 TU, respectively. The Stokes hydrodynamic radii of univalent and divalent carborane anions varied in the range 0.34-0.52 and 0.42-0.52 nm, respectively.
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- 2020
20. Druggable Hot Spots in the Schistosomiasis Cathepsin B1 Target Identified by Functional and Binding Mode Analysis of Potent Vinyl Sulfone Inhibitors
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William R. Roush, Michael Mareš, Pavla Fajtová, Jiří Brynda, Martin Lepšík, Cory D. Emal, Pavlína Řezáčová, Adam R. Renslo, Adéla Jílková, Jindřich Fanfrlík, Martin Horn, Helena Mertlíková-Kaiserová, Conor R. Caffrey, and Petra Rubešová
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0301 basic medicine ,Peptidomimetic ,cathepsin B ,030106 microbiology ,Druggability ,Schistosomiasis ,Cathepsin B ,Article ,drug target ,03 medical and health sciences ,medicine ,Moiety ,Animals ,Humans ,Sulfones ,vinyl sulfone inhibitor ,Schistosoma ,biology ,Chemistry ,Rational design ,Schistosoma mansoni ,biology.organism_classification ,medicine.disease ,030104 developmental biology ,Infectious Diseases ,Biochemistry ,parasite ,cysteine peptidase - Abstract
Schistosomiasis, a parasitic disease caused by blood flukes of the genus Schistosoma, is a global health problem with over 200 million people infected. Treatment relies on just one drug, and new chemotherapies are needed. Schistosoma mansoni cathepsin B1 (SmCB1) is a critical peptidase for the digestion of host blood proteins and a validated drug target. We screened a library of peptidomimetic vinyl sulfones against SmCB1 and identified the most potent SmCB1 inhibitors reported to date that are active in the subnanomolar range with second order rate constants (k2nd) of ∼2 × 105 M-1 s-1. High resolution crystal structures of the two best inhibitors in complex with SmCB1 were determined. Quantum chemical calculations of their respective binding modes identified critical hot spot interactions in the S1' and S2 subsites. The most potent inhibitor targets the S1' subsite with an N-hydroxysulfonic amide moiety and displays favorable functional properties, including bioactivity against the pathogen, selectivity for SmCB1 over human cathepsin B, and reasonable metabolic stability. Our results provide structural insights for the rational design of next-generation SmCB1 inhibitors as potential drugs to treat schistosomiasis.
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- 2020
21. The structural basis for the selectivity of sulfonamido dicarbaboranes toward cancer-associated carbonic anhydrase IX
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Jiří Brynda, Pavlína Řezáčová, Suzan El Anwar, K. Pospisilova, Josef Holub, Milan Fábry, Dmytro Bavol, Miroslav Havránek, Vlastimil Král, M. Kugler, and Bohumír Grüner
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Boron Compounds ,Carbonic anhydrase IX ,enzyme inhibitors ,Antineoplastic Agents ,RM1-950 ,Crystallography, X-Ray ,01 natural sciences ,carborane ,Antigens, Neoplasm ,Catalytic Domain ,Drug Discovery ,medicine ,Structure–activity relationship ,Humans ,Amino Acid Sequence ,Carbonic Anhydrase Inhibitors ,Pharmacology ,Solid tumour ,Sulfonamides ,010405 organic chemistry ,Chemistry ,structure-activity relationship ,Cancer ,A protein ,General Medicine ,Carbonic Anhydrase IX ,medicine.disease ,0104 chemical sciences ,010404 medicinal & biomolecular chemistry ,HEK293 Cells ,Biochemistry ,Carborane ,Therapeutics. Pharmacology ,Drug Screening Assays, Antitumor ,Selectivity ,Hydrophobic and Hydrophilic Interactions ,Protein Binding ,Research Article ,Research Paper - Abstract
Human carbonic anhydrase IX (CA IX), a protein specifically expressed on the surface of solid tumour cells, represents a validated target both for anticancer therapy and diagnostics. We recently identified sulfonamide dicarbaboranes as promising inhibitors of CA IX with favourable activities both in vitro and in vivo. To explain their selectivity and potency, we performed detailed X-ray structural analysis of their interactions within the active sites of CA IX and CA II. Series of compounds bearing various aliphatic linkers between the dicarbaborane cluster and sulfonamide group were examined. Preferential binding towards the hydrophobic part of the active site cavity was observed. Selectivity towards CA IX lies in the shape complementarity of the dicarbaborane cluster with a specific CA IX hydrophobic patch containing V131 residue. The bulky side chain of F131 residue in CA II alters the shape of the catalytic cavity, disrupting favourable interactions of the spherical dicarbaborane cluster.
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- 2020
22. Cobalt Bis(dicarbollide) Alkylsulfonamides: Potent and Highly Selective Inhibitors of Tumor Specific Carbonic Anhydrase IX
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Suzan El Anwar, K. Pospisilova, Josef Holub, Pavlína Řezáčová, Dmytro Bavol, Milan Fábry, Zdeňka Růžičková, Vlastimil Král, M. Kugler, Jiří Brynda, Bohumír Grüner, and Jan Nekvinda
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Sulfonamides ,Binding Sites ,010405 organic chemistry ,Stereochemistry ,Tumor specific ,Molecular Conformation ,chemistry.chemical_element ,General Chemistry ,Cobalt ,Carbonic Anhydrase IX ,010402 general chemistry ,Highly selective ,Crystallography, X-Ray ,01 natural sciences ,0104 chemical sciences ,Molecular Docking Simulation ,chemistry ,Coordination Complexes ,Catalytic Domain ,Neoplasms ,Humans ,Carbonic Anhydrase Inhibitors - Abstract
Carbonic anhydrase IX (CAIX) is an enzyme expressed on the surface of cells in hypoxic tumors. It plays a role in regulation of tumor pH and promotes thus tumor cell survival and occurrence of metastases. Here, derivatives of the cobalt bis(dicarbollide)(1-) anion are reported that are based on substitution at the carbon sites of the polyhedra by two alkylsulfonamide groups differing in the length of the aliphatic connector (from C1 to C4, n=1-4), which were prepared by cobalt insertion into the 7-sulfonamidoalkyl-7,8-dicarba-nido-undecaborate ions. Pure meso- and rac-diastereoisomeric forms were isolated. The series is complemented with monosubstituted species (n=2). Synthesis by a direct method furnished similar derivatives (n=2, 3), which are chlorinated at the B(8,8') boron sites. All compounds inhibited CAIX with subnanomolar inhibition constants and showed high selectivity for CAIX. The best inhibitory properties were observed for the compound with n= 3 and two substituents present in rac-arrangement with K
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- 2020
23. Sulfonamido carboranes as highly selective inhibitors of cancer-specific carbonic anhydrase IX
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Viswanath Das, Jan Nekvinda, Jiří Brynda, K. Pospisilova, Marian Hajduch, Stanislava Matějková, Jana Dvořanová, Martina Medvedíková, Josef Holub, Soňa Gurská, Bohumír Grüner, Miroslav Havránek, Vlastimil Král, M. Kugler, Barbora Liskova, Pavlína Řezáčová, Suzan El Anwar, Václav Šícha, Milan Fábry, and Petr Džubák
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Cell ,Antineoplastic Agents ,Breast Neoplasms ,Mice, SCID ,01 natural sciences ,03 medical and health sciences ,Mice ,Structure-Activity Relationship ,Dogs ,Antigens, Neoplasm ,Drug Discovery ,medicine ,Animals ,Humans ,Doxorubicin ,Cytotoxicity ,Carbonic Anhydrase IX ,Carbonic Anhydrase Inhibitors ,Cells, Cultured ,030304 developmental biology ,ADME ,Cell Proliferation ,Pharmacology ,chemistry.chemical_classification ,0303 health sciences ,Mice, Inbred BALB C ,Sulfonamides ,biology ,Dose-Response Relationship, Drug ,Molecular Structure ,010405 organic chemistry ,Organic Chemistry ,Active site ,General Medicine ,Neoplasms, Experimental ,Recombinant Proteins ,0104 chemical sciences ,Enzyme ,medicine.anatomical_structure ,chemistry ,Biochemistry ,Cell culture ,biology.protein ,Carborane ,Drug Screening Assays, Antitumor ,medicine.drug - Abstract
Carbonic anhydrase IX (CA IX) is a transmembrane enzyme overexpressed in hypoxic tumors, where it plays an important role in tumor progression. Specific CA IX inhibitors potentially could serve as anti-cancer drugs. We designed a series of sulfonamide inhibitors containing carborane clusters based on prior structural knowledge of carborane binding into the enzyme active site. Two types of carborane clusters, 12-vertex dicarba-closo-dodecaborane and 11-vertex 7,8-dicarba-nido-undecaborate (dicarbollide), were connected to a sulfonamide moiety via aliphatic linkers of varying lengths (1–4 carbon atoms; n = 1–4). In vitro testing of CA inhibitory potencies revealed that the optimal linker length for selective inhibition of CA IX was n = 3. A 1-sulfamidopropyl-1,2-dicarba-closo-dodecaborane (3) emerged as the strongest CA IX inhibitor from this series, with a Ki value of 0.5 nM and roughly 1230-fold selectivity towards CA IX over CA II. X-ray studies of 3 yielded structural insights into their binding modes within the CA IX active site. Compound 3 exhibited moderate cytotoxicity against cancer cell lines and primary cell lines in 2D cultures. Cytotoxicity towards multicellular spheroids was also observed. Moreover, 3 significantly lowered the amount of CA IX on the cell surface both in 2D cultures and spheroids and facilitated penetration of doxorubicin. Although 3 had only a moderate effect on tumor size in mice, we observed favorable ADME properties and pharmacokinetics in mice, and preferential presence in brain over serum.
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- 2020
24. DISC918836-Supplement – Supplemental material for Identification of Novel Carbonic Anhydrase IX Inhibitors Using High-Throughput Screening of Pooled Compound Libraries by DNA-Linked Inhibitor Antibody Assay (DIANA)
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Tykvart, Jan, Navrátil, Václav, Kugler, Michael, Šácha, Pavel, Jiří Schimer, Hlaváčková, Anna, Lukáš Tenora, Zemanová, Jitka, Dejmek, Milan, Král, Vlastimil, Potáček, Milan, Majer, Pavel, Jahn, Ullrich, Jiří Brynda, Řezáčová, Pavlína, and Konvalinka, Jan
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FOS: Clinical medicine ,111599 Pharmacology and Pharmaceutical Sciences not elsewhere classified - Abstract
Supplemental material, DISC918836-Supplement for Identification of Novel Carbonic Anhydrase IX Inhibitors Using High-Throughput Screening of Pooled Compound Libraries by DNA-Linked Inhibitor Antibody Assay (DIANA) by Jan Tykvart, Václav Navrátil, Michael Kugler, Pavel Šácha, Jiří Schimer, Anna Hlaváčková, Lukáš Tenora, Jitka Zemanová, Milan Dejmek, Vlastimil Král, Milan Potáček, Pavel Majer, Ullrich Jahn, Jiří Brynda, Pavlína Řezáčová and Jan Konvalinka in SLAS Discovery
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- 2020
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25. Crystal structure of native α-<scp>L</scp>-rhamnosidase from Aspergillus terreus
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Alena Křenková, Zbyszek Otwinowski, Petr Pachl, Pavel Mader, Jana Škerlová, Daniela Šimčíková, Vladimír Křen, Michael Kotik, Jiří Brynda, Pavlína Řezáčová, and Jana Kapešová
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Models, Molecular ,0301 basic medicine ,Glycosylation ,Glycoside Hydrolases ,Protein Conformation ,Stereochemistry ,030106 microbiology ,chemistry.chemical_element ,Crystallography, X-Ray ,03 medical and health sciences ,chemistry.chemical_compound ,Structural Biology ,Catalytic Domain ,Cleave ,Aspergillus terreus ,Alpha-L-rhamnosidase ,Growth medium ,biology ,Substrate (chemistry) ,Active site ,biology.organism_classification ,Sulfur ,Aspergillus ,chemistry ,biology.protein ,Function (biology) - Abstract
α-L-Rhamnosidases cleave terminal nonreducing α-L-rhamnosyl residues from many natural rhamnoglycosides. This makes them catalysts of interest for various biotechnological applications. The X-ray structure of the GH78 family α-L-rhamnosidase from Aspergillus terreus has been determined at 1.38 Å resolution using the sulfur single-wavelength anomalous dispersion phasing method. The protein was isolated from its natural source in the native glycosylated form, and the active site contained a glucose molecule, probably from the growth medium. In addition to its catalytic domain, the α-L-rhamnosidase from A. terreus contains four accessory domains of unknown function. The structural data suggest that two of these accessory domains, E and F, might play a role in stabilizing the aglycon portion of the bound substrate.
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- 2018
26. Enzymatic Preparation of 2'-5',3'-5'-Cyclic Dinucleotides, Their Binding Properties to Stimulator of Interferon Genes Adaptor Protein, and Structure/Activity Correlations
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Zahra Aliakbar Tehrani, Martin Zavřel, Tomáš Jandušík, Andrea Brázdová, Ondrej Gutten, Radim Nencka, Gabriel Birkus, Miloš Buděšínský, Ondřej Páv, Evžen Bouřa, Markéta Pimková Polidarová, Barbora Novotna, Radek Liboska, Zdeněk Vavřina, Milan Dejmek, Miroslav Smola, Jiří Brynda, Ivan Štěpánek, Lubomír Rulíšek, and Lenka Vaneková
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Chemistry ,Protein Conformation ,Signal transducing adaptor protein ,Membrane Proteins ,IκB kinase ,Cell biology ,Sting ,Structure-Activity Relationship ,HEK293 Cells ,Gene Expression Regulation ,Interferon ,Stimulator of interferon genes ,Drug Discovery ,Second messenger system ,medicine ,Leukocytes, Mononuclear ,Molecular Medicine ,Cytokines ,Humans ,Biological Assay ,Computer Simulation ,Nucleotides, Cyclic ,IRF3 ,Interferon regulatory factors ,medicine.drug - Abstract
Cyclic dinucleotides are second messengers in the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway, which plays an important role in recognizing tumor cells and viral or bacterial infections. They bind to the STING adaptor protein and trigger expression of cytokines via TANK binding kinase 1 (TBK1)/interferon regulatory factor 3 (IRF3) and inhibitor of nuclear factor-κB (IκB) kinase (IKK)/nuclear factor-κB (NFκB) signaling cascades. In this work, we describe an enzymatic preparation of 2'-5',3'-5'-cyclic dinucleotides (2'3'CDNs) with use of cyclic GMP-AMP synthases (cGAS) from human, mouse, and chicken. We profile substrate specificity of these enzymes by employing a small library of nucleotide-5'-triphosphate (NTP) analogues and use them to prepare 33 2'3'CDNs. We also determine affinity of these CDNs to five different STING haplotypes in cell-based and biochemical assays and describe properties needed for their optimal activity toward all STING haplotypes. Next, we study their effect on cytokine and chemokine induction by human peripheral blood mononuclear cells (PBMCs) and evaluate their cytotoxic effect on monocytes. Additionally, we report X-ray crystal structures of two new CDNs bound to STING protein and discuss structure-activity relationship by using quantum and molecular mechanical (QM/MM) computational modeling.
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- 2019
27. Structure-Based Optimization of Bisphosphonate Nucleoside Inhibitors of Human 5′(3′)-deoxyribonucleotidases
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Miloš Buděšínský, Ondřej Šimák, Petr Pachl, Pavlína Řezáčová, Jiří Brynda, and Ivan Rosenberg
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0301 basic medicine ,chemistry.chemical_classification ,Stereochemistry ,Chemistry ,medicine.medical_treatment ,Organic Chemistry ,Bisphosphonate ,03 medical and health sciences ,030104 developmental biology ,Enzyme ,Hydrolase ,medicine ,Structure based ,Physical and Theoretical Chemistry ,Nucleoside - Published
- 2018
28. Ranking Power of the SQM/COSMO Scoring Function on Carbonic Anhydrase II–Inhibitor Complexes
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Adam Pecina, Jiří Brynda, Lukáš Vrzal, Ramachandran Gnanasekaran, Magdalena Hořejší, Saltuk M. Eyrilmez, Jan Řezáč, Martin Lepšík, Pavlína Řezáčová, Pavel Hobza, Pavel Majer, Václav Veverka, and Jindřich Fanfrlík
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Sulfonamides ,010405 organic chemistry ,Ligands ,010402 general chemistry ,Carbonic Anhydrase II ,01 natural sciences ,Atomic and Molecular Physics, and Optics ,0104 chemical sciences ,Molecular Docking Simulation ,Models, Chemical ,Drug Design ,Quantum Theory ,Physical and Theoretical Chemistry ,Carbonic Anhydrase Inhibitors ,Protein Binding - Abstract
Accurate prediction of protein-ligand binding affinities is essential for hit-to-lead optimization and virtual screening. The reliability of scoring functions can be improved by including quantum effects. Here, we demonstrate the ranking power of the semiempirical quantum mechanics (SQM)/implicit solvent (COSMO) scoring function by using a challenging set of 10 inhibitors binding to carbonic anhydrase II through Zn
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- 2018
29. Fluidized Bed Incineration of Sewage Sludge in O2/N2 and O2/CO2 Atmospheres
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Siarhei Skoblia, Boleslav Zach, Jiří Brynda, Tomáš Durda, Michael Pohořelý, Karel Svoboda, Jaroslav Moško, Michal Jeremiáš, Šárka Václavková, Zdeněk Beňo, and Michal Šyc
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Pollutant ,Flue gas ,Waste management ,020209 energy ,General Chemical Engineering ,Energy Engineering and Power Technology ,02 engineering and technology ,Contamination ,Combustion ,Incineration ,Fuel Technology ,Volume (thermodynamics) ,Fluidized bed ,0202 electrical engineering, electronic engineering, information engineering ,Environmental science ,Sludge - Abstract
Sewage sludge incineration in a fluidized bed is considered to be one of the most suitable ways of sewage sludge disposal. This process reduces the volume of the waste and causes the destruction of organic contaminants such as POPs, pharmaceuticals and other compounds with endocrine-disrupting potential. Oxygen-enriched air combustion and oxy-fuel combustion can increase the combustion efficiency, reduce the amount of flue gas and make possible CO2 capture more effective. However, the influence of incineration medium composition has not yet been thoroughly investigated in case of sewage sludge incineration. In this paper, the incineration of sewage sludge in a bubbling fluidized bed reactor was studied at oxygen-enriched air conditions, oxy-fuel conditions and oxy-fuel conditions with zero and non-zero concentrations of steam, CO, NO, N2O and SO2 in the inlet combustion medium. Consequently, the effects of various operating parameters on pollutants formation were comprehensively described with emphasis on...
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- 2018
30. Structure-assisted design of inhibitors of CA IX enzyme based on polyhedral boron compounds
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Jiří Brynda, Michael Kugler, Jan Nekvinda, Josef Holub, Suzan El Anwar, Wiswanath Das, Václav Šícha, Klára Pospíšilová, Milan Fábry, Vlastimil Král, Petr Pachl, Marián Hajdúch, Bohumír Grüner, and Pavlína Řezáčová
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Inorganic Chemistry ,Structural Biology ,General Materials Science ,Physical and Theoretical Chemistry ,Condensed Matter Physics ,Biochemistry - Published
- 2021
31. Thermodynamic aspects of gasification derived syngas desulfurization, removal of hydrogen halides and regeneration of spent sorbents based on La2O3/La2O2CO3 and cerium oxides
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Yau-Pin Chyou, Miloslav Hartman, Michal Šyc, Michael Pohořelý, Jindřich Leitner, Jaromír Havlica, Jiří Brynda, Karel Svoboda, and Po-Chuang Chen
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Hydrogen ,Chemistry ,General Chemical Engineering ,Organic Chemistry ,Inorganic chemistry ,Energy Engineering and Power Technology ,chemistry.chemical_element ,Halide ,Producer gas ,02 engineering and technology ,010402 general chemistry ,021001 nanoscience & nanotechnology ,01 natural sciences ,Sulfur ,0104 chemical sciences ,Flue-gas desulfurization ,Gibbs free energy ,Cerium ,symbols.namesake ,Fuel Technology ,symbols ,0210 nano-technology ,Syngas - Abstract
The thermodynamic analyses are focused on the determination of theoretical equilibrium concentrations of H2S, HCl and HF for La2O3, La2O2CO3 and CexOy-based sulfur compounds sorbents, on possible interferences of hydrogen halides in H2S removal in the temperature range 600–1100 K and on upstream selective deep removal of HCl and HF in producer gas. Minimization of Gibbs energy for the system was used in more complicated situations. The lowest H2S concentration (
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- 2017
32. Rutinosidase from Aspergillus niger: crystal structure and insight into the enzymatic activity
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Jiří Brynda, Lada Biedermannová, Pavla Bojarová, Jana Kapešová, Pavlína Řezáčová, Vladimír Křen, Michael Kotik, Petr Pachl, and Helena Pelantová
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0301 basic medicine ,Models, Molecular ,Glycoside Hydrolases ,Stereochemistry ,Protein Conformation ,Rutin ,Protein Data Bank (RCSB PDB) ,Sequence Homology ,Crystallography, X-Ray ,Biochemistry ,Fungal Proteins ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,Catalytic Domain ,Hydrolase ,Amino Acid Sequence ,Molecular Biology ,chemistry.chemical_classification ,biology ,Active site ,Glycoside ,Glycosidic bond ,Cell Biology ,Rutinose ,030104 developmental biology ,chemistry ,030220 oncology & carcinogenesis ,Mutation ,biology.protein ,Aspergillus niger ,Quercetin ,Oxidation-Reduction - Abstract
Rutinosidases (α-l-rhamnosyl-β-d-glucosidases) catalyze the cleavage of the glycosidic bond between the aglycone and the disaccharide rutinose (α-l-rhamnopyranosyl-(1→6)-β-d-glucopyranose) of specific flavonoid glycosides such as rutin (quercetin 3-O-rutinoside). Microbial rutinosidases are part of the rutin catabolic pathway, enabling the microorganism to utilize rutin and related plant phenolic glycosides. Here, we report the first three-dimensional structure of a rutinosidase determined at 1.27-A resolution. The rutinosidase from Aspergillus niger K2 (AnRut), a member of glycoside hydrolase family GH-5, subfamily 23, was heterologously produced in Pichia pastoris. The X-ray structure of AnRut is represented by a distorted (β/α)8 barrel fold with its closest structural homologue being an exo-β-(1,3)-glucanase from Candida albicans (CaExg). The catalytic site is located in a deep pocket with a striking structural similarity to CaExg. However, the entrance to the active site of AnRut has been found to be different from that of CaExg - a mostly unstructured section of ~ 40 residues present in CaExg is missing in AnRut, whereas an additional loop of 13 amino acids partially covers the active site of AnRut. NMR analysis of reaction products provided clear evidence for a retaining reaction mechanism of AnRut. Unexpectedly, quercetin 3-O-glucoside was found to be a better substrate than rutin, and thus, AnRut cannot be considered a typical diglycosidase. Mutational analysis of conserved active site residues in combination with in silico modeling allowed identification of essential interactions for enzyme activity and helped to reveal further details of substrate binding. The protein sequence of AnRut has been revised. DATABASES: The nucleotide sequence of the rutinosidase-encoding gene is available in the GenBank database under the accession number MN393234. Structural data are available in the PDB database under the accession number 6I1A. ENZYME: α-l-Rhamnosyl-β-d-glucosidase (EC 3.2.1.168).
- Published
- 2019
33. Front Cover: Cobalt Bis(dicarbollide) Alkylsulfonamides: Potent and Highly Selective Inhibitors of Tumor Specific Carbonic Anhydrase IX (3/2021)
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Vlastimil Král, M. Kugler, Suzan El Anwar, Pavlína Řezáčová, Jiří Brynda, Milan Fábry, Zdeňka Růžičková, Josef Holub, Bohumír Grüner, Dmytro Bavol, Jan Nekvinda, and K. Pospisilova
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Front cover ,chemistry ,Tumor specific ,chemistry.chemical_element ,General Chemistry ,Carbonic Anhydrase IX ,Highly selective ,Cobalt ,Combinatorial chemistry - Published
- 2020
34. Unraveling the anti-influenza effect of flavonoids: Experimental validation of luteolin and its congeners as potent influenza endonuclease inhibitors
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Miroslav Flieger, Jan Weber, Jan Konvalinka, Ales Machara, Jindřich Fanfrlík, Pavel Majer, Jiří Brynda, Jan Hodek, Milan Kožíšek, Kateřina Radilová, Elena Karlukova, Václav Zima, and Carlos Berenguer Albiñana
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Drug Evaluation, Preclinical ,Microbial Sensitivity Tests ,Crystallography, X-Ray ,Antiviral Agents ,01 natural sciences ,Structure-Activity Relationship ,Viral Proteins ,03 medical and health sciences ,chemistry.chemical_compound ,Endonuclease ,Protein Domains ,RNA polymerase ,Drug Discovery ,Enzyme Inhibitors ,IC50 ,Enzyme Assays ,030304 developmental biology ,Flavonoids ,Pharmacology ,Orientin ,0303 health sciences ,Molecular Structure ,biology ,010405 organic chemistry ,Organic Chemistry ,Active site ,General Medicine ,Endonucleases ,RNA-Dependent RNA Polymerase ,0104 chemical sciences ,chemistry ,Biochemistry ,Influenza A virus ,biology.protein ,Neuraminidase ,Luteolin ,DNA ,Protein Binding - Abstract
The biological effects of flavonoids on mammal cells are diverse, ranging from scavenging free radicals and anti-cancer activity to anti-influenza activity. Despite appreciable effort to understand the anti-influenza activity of flavonoids, there is no clear consensus about their precise mode-of-action at a cellular level. Here, we report the development and validation of a screening assay based on AlphaScreen technology and illustrate its application for determination of the inhibitory potency of a large set of polyols against PA N-terminal domain (PA-Nter) of influenza RNA-dependent RNA polymerase featuring endonuclease activity. The most potent inhibitors we identified were luteolin with an IC50 of 72 ± 2 nM and its 8-C-glucoside orientin with an IC50 of 43 ± 2 nM. Submicromolar inhibitors were also evaluated by an in vitro endonuclease activity assay using single-stranded DNA, and the results were in full agreement with data from the competitive AlphaScreen assay. Using X-ray crystallography, we analyzed structures of the PA-Nter in complex with luteolin at 2.0 A resolution and quambalarine B at 2.5 A resolution, which clearly revealed the binding pose of these polyols coordinated to two manganese ions in the endonuclease active site. Using two distinct assays along with the structural work, we have presumably identified and characterized the molecular mode-of-action of flavonoids in influenza-infected cells.
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- 2020
35. Wood chips gasification in a fixed-bed multi-stage gasifier for decentralized high-efficiency CHP and biochar production: Long-term commercial operation
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Michal Šyc, Siarhei Skoblia, Michael Pohořelý, Lukáš Trakal, Michal Jeremiáš, Zdeněk Beňo, Karel Svoboda, Jaroslav Moško, Karel Soukup, Jiří Brynda, and Boleslav Zach
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Waste management ,Wood gas generator ,Power station ,business.industry ,020209 energy ,General Chemical Engineering ,Organic Chemistry ,Energy Engineering and Power Technology ,Tar ,Biomass ,Producer gas ,02 engineering and technology ,Renewable energy ,Fuel Technology ,020401 chemical engineering ,Biochar ,0202 electrical engineering, electronic engineering, information engineering ,Environmental science ,Char ,0204 chemical engineering ,business - Abstract
Gasification of woody biomass coupled with combined heat and power (CHP) production is an effective way to produce renewable energy from woody biomass. The tar content of the producer gas is considered to be the most problematic aspect of a biomass gasification for CHP production. Staged gasification allows the minimization of the tar production by the optimization of individual thermochemical subprocesses. This paper presents the operational results from a newly developed type of fixed-bed multi-stage gasifier GP750, which combines the principles of both twin-fire and two-stage gasification. This unique construction enables the production of a low-tar producer gas even by units with a power output as high as 0.7 MW. Here, we present the data measured during stable operation of the gasifier in a CHP plant operated by BOR Biotechnology company, including the composition of permanent gases measured on-line and the comprehensive gas and tar composition from the long-term operation, measured off-line. The measurements proved that the gasifier can produce a gas with low tar content (5–50 mg/m3), which is well beyond the limits for internal combustion engines. The energetic balance showed that the gross efficiency of the production of electricity of the whole power plant was as high as 32.9%. The gasifier also produced 36 kg of char per tonne of dry fuel on average. This char had a surface area in the range of 350–700 m2/g and very low content of both volatile matter and PAHs. Therefore, the char produced could potentially be sold as biochar.
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- 2020
36. Quantum Calculations Indicate Effective Electron Transfer between FMN and Benzoquinone in a New Crystal Structure of Escherichia coli WrbA
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Jannette Carey, Jiří Brynda, Michal Kuty, Ivana Kuta Smatanova, Rüdiger Ettrich, Dhiraj Sinha, Olga Ettrichova, Oksana Degtjarik, and David Řeha
- Subjects
0301 basic medicine ,Flavin Mononucleotide ,Crystal structure ,Crystallography, X-Ray ,010402 general chemistry ,Quinone oxidoreductase ,01 natural sciences ,Electron Transport ,03 medical and health sciences ,Electron transfer ,Oxidoreductase ,Computational chemistry ,Catalytic Domain ,Benzoquinones ,Escherichia coli ,Materials Chemistry ,Physical and Theoretical Chemistry ,Quantum ,chemistry.chemical_classification ,Escherichia coli Proteins ,Benzoquinone ,Protein Structure, Tertiary ,0104 chemical sciences ,Surfaces, Coatings and Films ,Repressor Proteins ,030104 developmental biology ,chemistry ,Docking (molecular) ,Quantum Theory ,Protein crystallization - Abstract
Quantum mechanical calculations using the Marcus equation are applied to compare the electron-transfer probability for two distinct crystal structures of the Escherichia coli protein WrbA, an FMN-dependent NAD(P)H:quinone oxidoreductase, with the bound substrate benzoquinone. The calculations indicate that the position of benzoquinone in a new structure reported here and solved at 1.33 A resolution is more likely to be relevant for the physiological reaction of WrbA than a previously reported crystal structure in which benzoquinone is shifted by ∼5 A. Because the true electron-acceptor substrate for WrbA is not yet known, the present results can serve to constrain computational docking attempts with potential substrates that may aid in identifying the natural substrate(s) and physiological role(s) of this enzyme. The approach used here highlights a role for quantum mechanical calculations in the interpretation of protein crystal structures.
- Published
- 2016
37. LIMITATIONS OF DRY FLUE GAS TREATMENT BY SODIUM BICARBONATE: THE INFLUENCE OF FLUE GAS COMPOSITION
- Author
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Jaroslav Moško, M. Punčochář, Jiří Brynda, Michal Šyc, Michael Pohořelý, Karel Svoboda, Boleslav Zach, and Šárka Václavková
- Subjects
Waste-to-energy ,Flue gas ,chemistry.chemical_compound ,Sodium bicarbonate ,chemistry ,Composition (visual arts) ,Pulp and paper industry - Published
- 2018
38. Structure-based design of a bisphosphonate 5′(3′)-deoxyribonucleotidase inhibitor
- Author
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Milan Fábry, Petr Pachl, Miloš Buděšínský, Ivan Rosenberg, Pavlína Řezáčová, Ondřej Šimák, and Jiří Brynda
- Subjects
Pharmacology ,chemistry.chemical_classification ,Nucleoside analogue ,Stereochemistry ,medicine.medical_treatment ,Organic Chemistry ,Phosphate ion ,Pharmaceutical Science ,Bisphosphonate ,Biochemistry ,chemistry.chemical_compound ,Enzyme ,chemistry ,Drug Discovery ,medicine ,Molecular Medicine ,Structure based ,Nucleotide ,Nucleoside ,Phosphomonoesters ,medicine.drug - Abstract
Cellular 5′-nucleotidases regulating nucleotide/nucleoside pools are capable of dephosphorylating phosphomonoesters of important nucleoside analogue drugs, thus decreasing their therapeutic efficacy. Based on previously known inhibitor–enzyme complex structures, we developed a promising inhibitor by mimicking the phosphate ion and achieved 50- and 100-fold increases in the inhibitory potency towards cdN and mdN, respectively. Crystal structures of both complexes showed major differences in the inhibitor binding mode towards both enzymes.
- Published
- 2015
39. Novel Structural Mechanism of Allosteric Regulation of Aspartic Peptidases via an Evolutionarily Conserved Exosite
- Author
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Ora Schueler-Furman, Martin Horn, Jiří Brynda, Jiří Vondrášek, Nawsad Alam, Petr Kopáček, Iva Hánová, Michael Mareš, Radka Houštecká, Daniel Sojka, and Lucie Marešová
- Subjects
0301 basic medicine ,030103 biophysics ,Proteolysis ,Clinical Biochemistry ,Allosteric regulation ,Cathepsin D ,Crystallography, X-Ray ,Ligands ,Biochemistry ,03 medical and health sciences ,Enzyme activator ,Ticks ,Allosteric Regulation ,Zymogen ,Catalytic Domain ,Drug Discovery ,Hydrolase ,medicine ,Animals ,Amino Acid Sequence ,Protein precursor ,Molecular Biology ,Pharmacology ,Enzyme Precursors ,biology ,medicine.diagnostic_test ,Active site ,Hydrogen-Ion Concentration ,Cell biology ,Enzyme Activation ,Kinetics ,030104 developmental biology ,biology.protein ,Molecular Medicine ,Peptides ,Sequence Alignment - Abstract
Summary Pepsin-family aspartic peptidases are biosynthesized as inactive zymogens in which the propeptide blocks the active site until its proteolytic removal upon enzyme activation. Here, we describe a novel dual regulatory function for the propeptide using a set of crystal structures of the parasite cathepsin D IrCD1. In the IrCD1 zymogen, intramolecular autoinhibition by the intact propeptide is mediated by an evolutionarily conserved exosite on the enzyme core. After activation, the mature enzyme employs the same exosite to rebind a small fragment derived from the cleaved propeptide. This fragment functions as an effective natural inhibitor of mature IrCD1 that operates in a pH-dependent manner through a unique allosteric inhibition mechanism. The study uncovers the propeptide-binding exosite as a target for the regulation of pepsin-family aspartic peptidases and defines the structural requirements for exosite inhibition.
- Published
- 2017
40. Crystal structure of native β-N-acetylhexosaminidase isolated from Aspergillus oryzae sheds light onto its substrate specificity, high stability, and regulation by propeptide
- Author
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Petr Pachl, Pavlína Řezáčová, Petr Man, Jan Bláha, Ondřej Vaněk, Jana Škerlová, Zbyszek Otwinowski, Petr Pompach, Jiří Brynda, Petr Novák, Kateřina Hofbauerová, and Zdeněk Kukačka
- Subjects
0301 basic medicine ,Models, Molecular ,Glycosylation ,Stereochemistry ,Protein subunit ,Aspergillus oryzae ,Protein Data Bank (RCSB PDB) ,G(M2) Ganglioside ,Chitobiose ,Crystallography, X-Ray ,Ligands ,Biochemistry ,Substrate Specificity ,Fungal Proteins ,03 medical and health sciences ,chemistry.chemical_compound ,Catalytic Domain ,Hydrolase ,Glycoside hydrolase ,Protein Interaction Domains and Motifs ,Amino Acid Sequence ,Molecular Biology ,Conserved Sequence ,chemistry.chemical_classification ,Enzyme Precursors ,Binding Sites ,030102 biochemistry & molecular biology ,biology ,Chemistry ,Protein Stability ,G(M2) Activator Protein ,Active site ,Cell Biology ,Protein engineering ,beta-N-Acetylhexosaminidases ,030104 developmental biology ,Enzyme ,Structural Homology, Protein ,biology.protein ,Dimerization ,Protein Processing, Post-Translational ,Sequence Alignment - Abstract
β-N-acetylhexosaminidase from the fungus Aspergillus oryzae is a secreted extracellular enzyme that cleaves chitobiose into constituent monosaccharides. It belongs to the GH 20 glycoside hydrolase family and consists of two N-glycosylated catalytic cores noncovalently associated with two 10-kDa O-glycosylated propeptides. We used X-ray diffraction and mass spectrometry to determine the structure of A. oryzae β-N-acetylhexosaminidase isolated from its natural source. The three-dimensional structure determined and refined to a resolution of 2.3 A revealed that this enzyme is active as a uniquely tight dimeric assembly further stabilized by N- and O-glycosylation. The propeptide from one subunit forms extensive noncovalent interactions with the catalytic core of the second subunit in the dimer, and this chain swap suggests the distinctive structural mechanism of the enzyme's activation. Unique structural features of β-N-acetylhexosaminidase from A. oryzae define a very stable and robust framework suitable for biotechnological applications. The crystal structure reported here provides structural insights into the enzyme architecture as well as the detailed configuration of the active site. These insights can be applied to rational enzyme engineering. DATABASE Structural data are available in the PDB database under the accession number 5OAR. ENZYME β-N-acetylhexosaminidase (EC 3.2.1.52).
- Published
- 2017
41. Structural Basis for Inhibition of Mycobacterial and Human Adenosine Kinase by 7-Substituted 7-(Het)aryl-7-deazaadenine Ribonucleosides
- Author
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Jindřich Fanfrlík, Radek Pohl, Jiřina Stolaříková, Petr Džubák, Petr Nauš, Petr Konečný, Jiří Dostál, Hana Dvořáková, Ivan Votruba, Marian Hajduch, Aleš Hnízda, Jiří Brynda, Michal Hocek, Helena Zábranská, Pavlína Řezáčová, Vaclav Veverka, Aurelie Bourderioux, Iva Pichová, Michal Dušek, and Jan Snášel
- Subjects
Protein Conformation ,medicine.drug_class ,Stereochemistry ,Antitubercular Agents ,Drug Evaluation, Preclinical ,Microbial Sensitivity Tests ,Adenosine kinase ,Crystallography, X-Ray ,Antimycobacterial ,Structure-Activity Relationship ,chemistry.chemical_compound ,Adenosine Triphosphate ,Drug Discovery ,medicine ,Humans ,Transferase ,Structure–activity relationship ,Binding site ,Adenosine Kinase ,Nuclear Magnetic Resonance, Biomolecular ,Binding Sites ,biology ,Chemistry ,Adenine ,Mycobacterium tuberculosis ,respiratory system ,bacterial infections and mycoses ,Adenosine ,3. Good health ,ADK ,Biochemistry ,biology.protein ,Molecular Medicine ,Ribonucleosides ,Adenosine triphosphate ,medicine.drug - Abstract
Adenosine kinase (ADK) from Mycobacterium tuberculosis (Mtb) was selected as a target for design of antimycobacterial nucleosides. Screening of 7-(het)aryl-7-deazaadenine ribonucleosides with Mtb and human (h) ADKs and testing with wild-type and drug-resistant Mtb strains identified specific inhibitors of Mtb ADK with micromolar antimycobacterial activity and low cytotoxicity. X-ray structures of complexes of Mtb and hADKs with 7-ethynyl-7-deazaadenosine showed differences in inhibitor interactions in the adenosine binding sites. 1D (1)H STD NMR experiments revealed that these inhibitors are readily accommodated into the ATP and adenosine binding sites of Mtb ADK, whereas they bind preferentially into the adenosine site of hADK. Occupation of the Mtb ADK ATP site with inhibitors and formation of catalytically less competent semiopen conformation of MtbADK after inhibitor binding in the adenosine site explain the lack of phosphorylation of 7-substituted-7-deazaadenosines. Semiempirical quantum mechanical analysis confirmed different affinity of nucleosides for the Mtb ADK adenosine and ATP sites.
- Published
- 2014
42. GS-8374, a Prototype Phosphonate-Containing Inhibitor of HIV-1 Protease, Effectively Inhibits Protease Mutants with Amino Acid Insertions
- Author
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Jan Konvalinka, Tomáš Cihlář, Dorien de Jong, Kirsten M. Stray, Jiří Brynda, Noortje M van Maarseveen, Milan Kožíšek, Pavlína Řezáčová, Monique Nijhuis, and Klára Grantz Šašková
- Subjects
Models, Molecular ,medicine.medical_treatment ,Immunology ,Mutant ,Organophosphonates ,HIV Infections ,Biology ,Crystallography, X-Ray ,Microbiology ,law.invention ,HIV Protease ,HIV-1 protease ,law ,Virology ,Drug Resistance, Viral ,Vaccines and Antiviral Agents ,medicine ,Humans ,HIV Protease Inhibitor ,Amino Acid Sequence ,Binding site ,Peptide sequence ,chemistry.chemical_classification ,Binding Sites ,Protease ,virus diseases ,HIV Protease Inhibitors ,Molecular biology ,Amino acid ,Mutagenesis, Insertional ,chemistry ,Insect Science ,HIV-1 ,biology.protein ,Recombinant DNA - Abstract
Insertions in the protease (PR) region of human immunodeficiency virus (HIV) represent an interesting mechanism of antiviral resistance against HIV PR inhibitors (PIs). Here, we demonstrate the improved ability of a phosphonate-containing experimental HIV PI, GS-8374, relative to that of other PIs, to effectively inhibit patient-derived recombinant HIV strains bearing PR insertions and numerous other mutations. We correlate enzyme inhibition with the catalytic activities of corresponding recombinant PRs in vitro and provide a biochemical and structural analysis of the PR-inhibitor complex.
- Published
- 2014
43. Thermodynamic and structural analysis of HIV protease resistance to darunavir - analysis of heavily mutated patient-derived HIV-1 proteases
- Author
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Klára Grantz Šašková, Martin Lepšík, Pavlína Řezáčová, Jan Konvalinka, Milan Kožíšek, and Jiří Brynda
- Subjects
Proteases ,Stereochemistry ,medicine.medical_treatment ,Molecular Sequence Data ,Biochemistry ,HIV Protease ,Drug Resistance, Viral ,medicine ,HIV Protease Inhibitor ,Amino Acid Sequence ,Molecular Biology ,Darunavir ,Sulfonamides ,Protease ,Chemistry ,Isothermal titration calorimetry ,HIV Protease Inhibitors ,Cell Biology ,computer.file_format ,Resistance mutation ,Protein Data Bank ,Molecular Docking Simulation ,Mutation ,Thermodynamics ,computer ,Protein Binding ,Entropy (order and disorder) ,medicine.drug - Abstract
We report enzymologic, thermodynamic and structural analyses of a series of six clinically derived mutant HIV proteases (PR) resistant to darunavir. As many as 20 mutations in the resistant PRs decreased the binding affinity of darunavir by up to 13 000-fold, mostly because of a less favorable enthalpy of binding that was only partially compensated by the entropic contribution. X-ray structure analysis suggested that the drop in enthalpy of darunavir binding to resistant PR species was mostly the result of a decrease in the number of hydrogen bonds and a loosening of the fit between the inhibitor and the mutated enzymes. The favorable entropic contribution to darunavir binding to mutated PR variants correlated with a larger burial of the nonpolar solvent-accessible surface area upon inhibitor binding. We show that even very dramatic changes in the PR sequence leading to the loss of hydrogen bonds with the inhibitor could be partially compensated by the entropy contribution as a result of the burial of the larger nonpolar surface area of the mutated HIV PRs. Database Atomic coordinates and structure factors for the crystal structures PRwt–DRV and PRDRV2–DRV complex have been deposited in the Protein Data Bank under accession codes 4LL3 and 3TTP, respectively. Structured digital abstract • PR and PR bind by x-ray crystallography (View interaction)
- Published
- 2014
44. Structures of human cytosolic and mitochondrial nucleotidases: implications for structure-based design of selective inhibitors
- Author
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Milan Fábry, Pavlina Rezacova, Ondřej Šimák, Ivan Rosenberg, Jiří Brynda, and Petr Pachl
- Subjects
Models, Molecular ,Deoxyribonucleosides ,Protein Conformation ,Stereochemistry ,Deoxyribonucleotides ,Organophosphonates ,Protonation ,Crystallography, X-Ray ,Phosphates ,Structure-Activity Relationship ,Nucleotidases ,chemistry.chemical_compound ,Cytosol ,Structural Biology ,Catalytic Domain ,Hydrolase ,Escherichia coli ,Humans ,Nucleotide ,Enzyme Inhibitors ,chemistry.chemical_classification ,biology ,Chemistry ,Active site ,General Medicine ,Recombinant Proteins ,Mitochondria ,Isoenzymes ,Molecular Docking Simulation ,Deoxyribonucleoside ,Eukaryotic Cells ,Organ Specificity ,Drug Design ,biology.protein - Abstract
The human 5′(3′)-deoxyribonucleotidases catalyze the dephosphorylation of deoxyribonucleoside monophosphates to the corresponding deoxyribonucleosides and thus help to maintain the balance between pools of nucleosides and nucleotides. Here, the structures of human cytosolic deoxyribonucleotidase (cdN) at atomic resolution (1.08 Å) and mitochondrial deoxyribonucleotidase (mdN) at near-atomic resolution (1.4 Å) are reported. The attainment of an atomic resolution structure allowed interatomic distances to be used to assess the probable protonation state of the phosphate anion and the side chains in the enzyme active site. A detailed comparison of the cdN and mdN active sites allowed the design of a cdN-specific inhibitor.
- Published
- 2014
45. Backbone resonance assignments of human cytosolic dNT-1 nucleotidase
- Author
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Aleš Hnízda, Petr Pachl, Milan Fábry, Zdeněk Tošner, Radka Skleničková, Vaclav Veverka, and Jiří Brynda
- Subjects
chemistry.chemical_classification ,Nucleotidase activity ,Chemistry ,Biochemistry ,Small molecule ,Deoxyribonucleotides ,5'-nucleotidase ,Dephosphorylation ,Nucleotidases ,Structural Biology ,Drug Design ,Nucleotidase ,Humans ,Nucleotide ,Enzyme Inhibitors ,Nuclear Magnetic Resonance, Biomolecular ,Nucleoside - Abstract
Cytosolic dNT-1 nucleotidase plays a key role in the homeostasis of pyrimidine deoxyribonucleotides in mammalian cells. The enzyme is responsible for the dephosphorylation of physiological substrates as well as nucleoside analogues that are used in antiviral and anticancer therapies, therefore selective inhibition of the dNT-1 nucleotidase activity may lead to an increase in efficacy of this type of therapeutic compounds. Here, we report the backbone ¹H, ¹³C and ¹⁵N assignments for the 47 kDa dNT-1 dimer, which will be used for structural characterisation of dNT-1 complexes with small molecule inhibitors obtained through modification of pyrimidine nucleotide scaffolds or optimisation of successful binders obtained from the screening of fragment libraries.
- Published
- 2013
46. Carborane-Based Carbonic Anhydrase Inhibitors
- Author
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Mario Bakardiev, Václav Šícha, Pavel Mader, Milan Fábry, Petr Cigler, Pavlína Řezáčová, Bohumír Grüner, Kristýna Poncová, and Jiří Brynda
- Subjects
biology ,Drug discovery ,High selectivity ,Active site ,General Chemistry ,Crystal structure ,General Medicine ,Combinatorial chemistry ,Catalysis ,chemistry.chemical_compound ,Structure-Activity Relationship ,chemistry ,Carbonic anhydrase ,Drug Design ,Drug Discovery ,biology.protein ,Organic chemistry ,Carborane ,Humans ,Carbonic Anhydrase Inhibitors ,Sulfamide - Abstract
CA inhibitors: Human carbonic anhydrases (CAs) are diagnostic and therapeutic targets. Various carborane cages are shown to act as active-site-directed inhibitors, and substitution with a sulfamide group and other substituents leads to compounds with high selectivity towards the cancer-specific isozyme IX. Crystal structures of the carboranes in the active site provide information that can be applied to the structure-based design of specific inhibitors.
- Published
- 2013
47. Synthesis and biological evaluation of guanidino analogues of roscovitine
- Author
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Jiří Brynda, Zlatko Janeba, Iva Dolečková, Vladimír Kryštof, Jiří Voller, Martin Dračínský, and Michal Česnek
- Subjects
Models, Molecular ,Purine ,Cyclin E ,Stereochemistry ,Antineoplastic Agents ,Structure-Activity Relationship ,chemistry.chemical_compound ,Cyclin-dependent kinase ,CDC2 Protein Kinase ,Drug Discovery ,Humans ,Protein kinase A ,Cytotoxicity ,Protein Kinase Inhibitors ,Guanidine ,Cell Proliferation ,Pharmacology ,Cyclin-dependent kinase 1 ,Dose-Response Relationship, Drug ,Molecular Structure ,biology ,Kinase ,Chemistry ,Cell Cycle ,Cyclin-Dependent Kinase 2 ,Organic Chemistry ,Cyclin-dependent kinase 2 ,General Medicine ,Biochemistry ,Purines ,MCF-7 Cells ,biology.protein ,Drug Screening Assays, Antitumor - Abstract
A series of 2,9-substituted 6-guanidinopurines, structurally related to the cyclin-dependent kinase (CDK) inhibitors olomoucine and roscovitine, has been synthesized and characterized. A new copper-catalyzed method for the synthesis of 2-substituted 6-guanidino-9-isopropylpurines under mild reaction conditions has been developed. All prepared compounds were screened for their CDK1 and CDK2 inhibitory activities, cytotoxicity and antiproliferative effects in the breast cancer-derived cell line MCF7. The most active derivative 16g possessed an identical side chain in the C2 position to roscovitine; this compound displayed approximately five fold higher inhibitory activity towards CDK2/cyclin E and more than ten fold increase in cytotoxicity in MCF7 cells. Interestingly and in contrast to previously described findings, (S)-6-guanidinopurine derivatives were generally more active than their (R)-counterparts. Kinase selectivity profiling of (R)- and (S)-enantiomers 16e and 16g, respectively, revealed that introduction of a guanidino group at the C6 position of the purine moiety decreased selectivity towards protein kinases compared to roscovitine. Nevertheless, increased inhibitory activity and decreased selectivity offer a good starting point for further development of new protein kinase inhibitors.
- Published
- 2013
48. MOESM1 of Myristoylation drives dimerization of matrix protein from mouse mammary tumor virus
- Author
-
Doležal, Michal, Aleš Zábranský, Jiří Dostál, Ondřej Vaněk, Jiří Brynda, Lepšík, Martin, Hadravová, Romana, and Pichová, Iva
- Abstract
Additional file 1: Figure S1. MS MALDI TOF/TOF analysis of myristoylated, myr(+), and nonmyristoylated, myr(−), MMTV MA. (A) The sample of myristoylated MA contained myr(+) MA (m/z = 13006). The sample of nonmyristoylated MA contained myr(−) MA (m/z = 12795) and MA with uncleaved initial methionine (m/z = 12927). (B) To approximately determine the minimal detectable amount of myr(−) MA in the sample of myr(+) MA, the sample of myr(+) MA was mixed with the sample of myr(−) in the ratio 1000:1 (2:0.002 mg/mL). This amount of myr(−) MA was detectable whereas the sample of myr(+) contained no detectable amount of myr(−) MA.
- Published
- 2016
- Full Text
- View/download PDF
49. The crystal structure of protease Sapp1p from Candida parapsilosis in complex with the HIV protease inhibitor ritonavir
- Author
-
Jiří Dostál, Petr Pachl, Iva Pichová, Jiří Brynda, Pavlina Rezacova, and Olga Hrušková-Heidingsfeldová
- Subjects
Models, Molecular ,medicine.medical_treatment ,Crystallography, X-Ray ,Candida parapsilosis ,Virulence factor ,Microbiology ,Fungal Proteins ,Structure-Activity Relationship ,chemistry.chemical_compound ,Drug Discovery ,medicine ,Aspartic Acid Endopeptidases ,HIV Protease Inhibitor ,Candida ,Pharmacology ,chemistry.chemical_classification ,Ritonavir ,Protease ,biology ,Proteolytic enzymes ,HIV Protease Inhibitors ,General Medicine ,biology.organism_classification ,Enzyme ,Biochemistry ,chemistry ,Pepstatin ,medicine.drug - Abstract
Secreted aspartic proteases (Saps) are extracellular proteolytic enzymes that enhance the virulence of Candida pathogens. These enzymes therefore represent possible targets for therapeutic drug design. Saps are inhibited by nanomolar concentrations of the classical inhibitor of aspartic proteases pepstatin A and also by the inhibitors of the HIV protease, but with the K(i) of micromolar values or higher. To contribute to the discussion regarding whether HIV protease inhibitors can act against opportunistic mycoses by the inhibition of Saps, we determined the structure of Sapp1p from Candida parapsilosis in complex with ritonavir (RTV), a clinically used inhibitor of the HIV protease. The crystal structure refined at resolution 2.4 Å proved binding of RTV into the active site of Sapp1p and provided the structural information necessary to evaluate the stability and specificity of the protein-inhibitor interaction.
- Published
- 2011
50. Structural Basis for Inhibition of Cathepsin B Drug Target from the Human Blood Fluke, Schistosoma mansoni
- Author
-
Jiří Brynda, Conor R. Caffrey, Pavlína Řezáčová, Martin Horn, Jana Váchová, Adéla Jílková, James H. McKerrow, Jindřich Fanfrlík, Martin Lepšík, and Michael Mareš
- Subjects
Peptidomimetic ,Cathepsin D ,Biology ,Crystallography, X-Ray ,Biochemistry ,Cathepsin B ,Hemoglobins ,Structure-Activity Relationship ,Drug Delivery Systems ,Animals ,Humans ,Structure–activity relationship ,Protease Inhibitors ,Molecular Biology ,Schistosoma ,Hydrolysis ,Helminth Proteins ,Schistosoma mansoni ,Cell Biology ,biology.organism_classification ,Cysteine protease ,Molecular biology ,Blood proteins ,Schistosomiasis mansoni ,Protein Structure and Folding ,Peptidomimetics ,Peptides - Abstract
Schistosomiasis caused by a parasitic blood fluke of the genus Schistosoma afflicts over 200 million people worldwide. Schistosoma mansoni cathepsin B1 (SmCB1) is a gut-associated peptidase that digests host blood proteins as a source of nutrients. It is under investigation as a drug target. To further this goal, we report three crystal structures of SmCB1 complexed with peptidomimetic inhibitors as follows: the epoxide CA074 at 1.3 Å resolution and the vinyl sulfones K11017 and K11777 at 1.8 and 2.5 Å resolutions, respectively. Interactions of the inhibitors with the subsites of the active-site cleft were evaluated by quantum chemical calculations. These data and inhibition profiling with a panel of vinyl sulfone derivatives identify key binding interactions and provide insight into the specificity of SmCB1 inhibition. Furthermore, hydrolysis profiling of SmCB1 using synthetic peptides and the natural substrate hemoglobin revealed that carboxydipeptidase activity predominates over endopeptidolysis, thereby demonstrating the contribution of the occluding loop that restricts access to the active-site cleft. Critically, the severity of phenotypes induced in the parasite by vinyl sulfone inhibitors correlated with enzyme inhibition, providing support that SmCB1 is a valuable drug target. The present structure and inhibitor interaction data provide a footing for the rational design of anti-schistosomal inhibitors.
- Published
- 2011
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