40 results on '"Denys Pogoryelov"'
Search Results
2. Crystal structure of the translation recovery factor Trf from Sulfolobus solfataricus
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Marco Kaiser, Jan Philip Wurm, Birgit Märtens, Udo Bläsi, Denys Pogoryelov, and Jens Wöhnert
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DUF35 ,ribosome ,Sulfolobus solfataricus ,translation initiation ,translation recovery factor Trf ,Biology (General) ,QH301-705.5 - Abstract
During translation initiation, the heterotrimeric archaeal translation initiation factor 2 (aIF2) recruits the initiator tRNAi to the small ribosomal subunit. In the stationary growth phase and/or during nutrient stress, Sulfolobus solfataricus aIF2 has a second function: It protects leaderless mRNAs against degradation by binding to their 5′‐ends. The S. solfataricus protein Sso2509 is a translation recovery factor (Trf) that interacts with aIF2 and is responsible for the release of aIF2 from bound mRNAs, thereby enabling translation re‐initiation. It is a member of the domain of unknown function 35 (DUF35) protein family and is conserved in Sulfolobales as well as in other archaea. Here, we present the X‐ray structure of S. solfataricus Trf solved to a resolution of 1.65 Å. Trf is composed of an N‐terminal rubredoxin‐like domain containing a bound zinc ion and a C‐terminal oligosaccharide/oligonucleotide binding fold domain. The Trf structure reveals putative mRNA binding sites in both domains. Surprisingly, the Trf protein is structurally but not sequentially very similar to proteins linked to acyl‐CoA utilization—for example, the Sso2064 protein from S. solfataricus—as well as to scaffold proteins found in the acetoacetyl‐CoA thiolase/high‐mobility group‐CoA synthase complex of the archaeon Methanothermococcus thermolithotrophicus and in a steroid side‐chain‐cleaving aldolase complex from the bacterium Thermomonospora curvata. This suggests that members of the DUF35 protein family are able to act as scaffolding and binding proteins in a wide variety of biological processes.
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- 2020
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3. PENG: A Neural Gas-Based Approach for Pharmacophore Elucidation. Method Design, Validation, and Virtual Screening for Novel Ligands of LTA4H.
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Daniel Moser, Sandra K. Wittmann, Jan Kramer, René Blöcher, Janosch Achenbach, Denys Pogoryelov, and Ewgenij Proschak
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- 2015
- Full Text
- View/download PDF
4. Design of Dual Inhibitors of Soluble Epoxide Hydrolase and LTA4 Hydrolase
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René Blöcher, Kerstin Hiesinger, Sandra K. Wittmann, Jana Gerstmeier, Denys Pogoryelov, Annika Schott, Ewgenij Proschak, Finja Witt, Dieter Steinhilber, Oliver Werz, and Jan S. Kramer
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Epoxide hydrolase 2 ,010405 organic chemistry ,Organic Chemistry ,Dual inhibitor ,01 natural sciences ,Biochemistry ,0104 chemical sciences ,010404 medicinal & biomolecular chemistry ,chemistry.chemical_compound ,chemistry ,LTA4 hydrolase ,Drug Discovery ,cardiovascular system ,Arachidonic acid ,Polypharmacology - Abstract
Multitarget anti-inflammatory drugs interfering with the arachidonic acid cascade exhibit superior efficacy. In this study, a prototype dual inhibitor of soluble epoxide hydrolase (sEH) and LTA4 hy...
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- 2019
5. Discovery of the First in Vivo Active Inhibitors of the Soluble Epoxide Hydrolase Phosphatase Domain
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Stefano Woltersdorf, Thomas Duflot, Christophe Morisseau, Victor Hernandez-Olmos, Ewgenij Proschak, Apirat Chaikuad, Franca-M Klingler, Sandra K. Wittmann, Felix Knöll, Dieter Steinhilber, Jan S. Kramer, Felix F Lillich, Angelo Sala, Kerstin Hiesinger, Daniel Merk, Sylvain Fraineau, Jeremy Bellien, Bruce D. Hammock, Jan Heering, Steffen Brunst, Stefan Knapp, G. Enrico Rovati, Julie Rondeaux, Denys Pogoryelov, Carola Buccellati, Matthieu Leuillier, Fraineau, Sylvain, Goethe-University Frankfurt am Main, Endothélium, valvulopathies et insuffisance cardiaque (EnVI), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Rouen, Normandie Université (NU), University of California [Davis] (UC Davis), University of California (UC), Università degli Studi di Milano = University of Milan (UNIMI), Fraunhofer Institute for Molecular Biology and Applied Ecology (Fraunhofer IME), Fraunhofer (Fraunhofer-Gesellschaft), Publica, University of California, and Università degli Studi di Milano [Milano] (UNIMI)
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Male ,Epoxide hydrolase 2 ,[SDV.BIO]Life Sciences [q-bio]/Biotechnology ,Structure analysis ,Medicinal & Biomolecular Chemistry ,[SDV]Life Sciences [q-bio] ,Phosphatase ,Ligands ,01 natural sciences ,Article ,Rats, Sprague-Dawley ,Medicinal and Biomolecular Chemistry ,Structure-Activity Relationship ,03 medical and health sciences ,In vivo ,Catalytic Domain ,Drug Discovery ,Hydrolase ,Animals ,Humans ,Structure–activity relationship ,Phosphofructokinase 2 ,Enzyme Inhibitors ,Binding site ,Oxazoles ,030304 developmental biology ,Epoxide Hydrolases ,0303 health sciences ,Binding Sites ,Chemistry ,Organic Chemistry ,Temperature ,Pharmacology and Pharmaceutical Sciences ,Phosphoric Monoester Hydrolases ,[SDV.BIO] Life Sciences [q-bio]/Biotechnology ,Rats ,3. Good health ,0104 chemical sciences ,[SDV] Life Sciences [q-bio] ,010404 medicinal & biomolecular chemistry ,Biochemistry ,Drug Design ,cardiovascular system ,Molecular Medicine ,Sprague-Dawley - Abstract
International audience; The emerging pharmacological target soluble epoxide hydrolase (sEH) is a bifunctional enzyme exhibiting two different catalytic activities that are located in two distinct domains. Although the physiological role of the C-terminal hydrolase domain is well-investigated, little is known about its phosphatase activity, located in the N-terminal phosphatase domain of sEH (sEH-P). Herein we report the discovery and optimization of the first inhibitor of human and rat sEH-P that is applicable in vivo. X-ray structure analysis of the sEH phosphatase domain complexed with an inhibitor provides insights in the molecular basis of small-molecule sEH-P inhibition and helps to rationalize the structure−activity relationships. 4-(4-(3,4-Dichlorophenyl)-5-phenyloxazol-2-yl)butanoic acid (22b, SWE101) has an excellent pharmacokinetic and pharmacodynamic profile in rats and enables the investigation of the physiological and pathophysiological role of sEH-P in vivo.
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- 2019
6. Design, Synthesis, and Structure-Activity Relationship Studies of Dual Inhibitors of Soluble Epoxide Hydrolase and 5-Lipoxygenase
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Bettina Hofmann, Kerstin Hiesinger, Steffen Brunst, Timon Eckes, Carlo Angioni, Ewgenij Proschak, Dieter Steinhilber, Jan S. Kramer, Manfred Schubert-Zsilavecz, Gerd Geisslinger, Achim Schmidtko, Josef Pfeilschifter, Simon B.M. Kretschmer, Lilia Weizel, Sandra K. Wittmann, Sven George, Stephanie Schwalm, Sandra Beyer, Jan Heering, Cathrin Flauaus, Astrid Kaiser, and Denys Pogoryelov
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Epoxide hydrolase 2 ,Stereochemistry ,Neutrophils ,01 natural sciences ,03 medical and health sciences ,chemistry.chemical_compound ,Structure-Activity Relationship ,Drug Discovery ,Structure–activity relationship ,Animals ,Humans ,Lipoxygenase Inhibitors ,Cells, Cultured ,030304 developmental biology ,chemistry.chemical_classification ,Epoxide Hydrolases ,0303 health sciences ,Arachidonate 5-Lipoxygenase ,biology ,Molecular Structure ,Chemistry ,Anti-Inflammatory Agents, Non-Steroidal ,0104 chemical sciences ,Rats ,010404 medicinal & biomolecular chemistry ,Enzyme ,Design synthesis ,Drug Design ,Arachidonate 5-lipoxygenase ,cardiovascular system ,biology.protein ,Microsomes, Liver ,Molecular Medicine ,Arachidonic acid ,Pharmacophore ,Protein Binding - Abstract
Inhibition of multiple enzymes of the arachidonic acid cascade leads to synergistic anti-inflammatory effects. Merging of 5-lipoxygenase (5-LOX) and soluble epoxide hydrolase (sEH) pharmacophores led to the discovery of a dual 5-LOX/sEH inhibitor, which was subsequently optimized in terms of potency toward both targets and metabolic stability. The optimized lead structure displayed cellular activity in human polymorphonuclear leukocytes, oral bioavailability, and target engagement in vivo and demonstrated profound anti-inflammatory and anti-fibrotic efficiency in a kidney injury model caused by unilateral ureteral obstruction in mice. These results pave the way for investigating the therapeutic potential of dual 5-LOX/sEH inhibitors in other inflammation- and fibrosis-related disease models.
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- 2020
7. Cyclophilin anaCyp40 regulates photosystem assembly and phycobilisome association in a cyanobacterium
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Shivam, Yadav, Martin, Centola, Mathilda, Glaesmann, Denys, Pogoryelov, Roman, Ladig, Mike, Heilemann, L C, Rai, Özkan, Yildiz, and Enrico, Schleiff
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Cyclophilins ,Phycobilisomes ,Humans ,Photosystem II Protein Complex ,Cyanobacteria ,Thylakoids - Abstract
Cyclophilins, or immunophilins, are proteins found in many organisms including bacteria, plants and humans. Most of them display peptidyl-prolyl cis-trans isomerase activity, and play roles as chaperones or in signal transduction. Here, we show that cyclophilin anaCyp40 from the cyanobacterium Anabaena sp. PCC 7120 is enzymatically active, and seems to be involved in general stress responses and in assembly of photosynthetic complexes. The protein is associated with the thylakoid membrane and interacts with phycobilisome and photosystem components. Knockdown of anacyp40 leads to growth defects under high-salt and high-light conditions, and reduced energy transfer from phycobilisomes to photosystems. Elucidation of the anaCyp40 crystal structure at 1.2-Å resolution reveals an N-terminal helical domain with similarity to PsbQ components of plant photosystem II, and a C-terminal cyclophilin domain with a substrate-binding site. The anaCyp40 structure is distinct from that of other multi-domain cyclophilins (such as Arabidopsis thaliana Cyp38), and presents features that are absent in single-domain cyclophilins.
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- 2020
8. Challenges in the Development of a Thiol-Based Broad-Spectrum Inhibitor for Metallo-β-Lactamases
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Daniel Kohnhäuser, Markus Hartmann, Denia Frank, Sandra K. Wittmann, Dominik Büttner, Thomas A. Wichelhaus, Jan S. Kramer, Denys Pogoryelov, Christian Kurz, Lilia Weizel, Franca Maria Klingler, Astrid Brüggerhoff, Dieter Steinhilber, and Ewgenij Proschak
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0301 basic medicine ,chemistry.chemical_classification ,medicine.drug_class ,Antibiotics ,chemical and pharmacologic phenomena ,Biology ,bacterial infections and mycoses ,beta-Lactam Resistance ,In vitro ,03 medical and health sciences ,030104 developmental biology ,Infectious Diseases ,Enzyme ,Antibiotic resistance ,chemistry ,Biochemistry ,Drug Discovery ,Hydrolase ,Thiol ,medicine ,Moiety ,Sulfhydryl Compounds ,beta-Lactamase Inhibitors ,Antibacterial activity - Abstract
Pathogens, expressing metallo-β-lactamases (MBLs), become resistant against most β-lactam antibiotics. Besides the dragging search for new antibiotics, development of MBL inhibitors would be an alternative weapon against resistant bacterial pathogens. Inhibition of resistance enzymes could restore the antibacterial activity of β-lactams. Various approaches to MBL inhibitors are described; among others, the promising motif of a zinc coordinating thiol moiety is very popular. Nevertheless, since the first report of a thiol-based MBL inhibitor (thiomandelic acid) in 2001, no steps in development of thiol based MBL inhibitors were reported that go beyond clinical isolate testing. In this study, we report on the synthesis and biochemical characterization of thiol-based MBL inhibitors and highlight the challenges behind the development of thiol-based compounds, which exhibit good in vitro activity toward a broad spectrum of MBLs, selectivity against human off-targets, and reasonable activity against clinical isolates.
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- 2017
9. Thermodynamic properties of leukotriene A 4 hydrolase inhibitors
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René Bloecher, Lena Kalinowsky, Dieter Steinhilber, Jan S. Kramer, Stefan Knapp, Ewgenij Proschak, Jan Heering, Denys Pogoryelov, and Sandra K. Wittmann
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0301 basic medicine ,030102 biochemistry & molecular biology ,Stereochemistry ,Leukotriene B4 ,Leukotriene A4 ,In silico ,Organic Chemistry ,Clinical Biochemistry ,Hydrolase activity ,Pharmaceutical Science ,Biochemistry ,Leukotriene-A4 hydrolase ,03 medical and health sciences ,chemistry.chemical_compound ,030104 developmental biology ,chemistry ,Drug Discovery ,Hydrolase ,Molecular Medicine ,Phosphofructokinase 2 ,Binding site ,Molecular Biology - Abstract
The leukotriene A4 hydrolase (LTA4H) is a bifunctional enzyme, containing a peptidase and a hydrolase activity both activities having opposing functions regulating inflammatory response. The hydrolase activity is responsible for the conversion of leukotriene A4 to pro-inflammatory leukotriene B4, and hence, selective inhibitors of the hydrolase activity are of high pharmacological interest. Here we present the thermodynamic characterization of structurally distinct inhibitors of the LTA4H that occupy different regions of the binding site using different biophysical methods. An in silico method for the determination of stabilized water molecules in the binding site of the apo structure of LTA4H is used to interpret the measured thermodynamic data and provided insights for design of novel LTA4H inhibitors.
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- 2016
10. Design of Dual Inhibitors of Soluble Epoxide Hydrolase and LTA
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Kerstin, Hiesinger, Annika, Schott, Jan S, Kramer, René, Blöcher, Finja, Witt, Sandra K, Wittmann, Dieter, Steinhilber, Denys, Pogoryelov, Jana, Gerstmeier, Oliver, Werz, and Ewgenij, Proschak
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lipids (amino acids, peptides, and proteins) - Abstract
[Image: see text] Multitarget anti-inflammatory drugs interfering with the arachidonic acid cascade exhibit superior efficacy. In this study, a prototype dual inhibitor of soluble epoxide hydrolase (sEH) and LTA(4) hydrolase (LTA(4)H) with submicromolar activity toward both targets has been designed and synthesized. Preliminary structure–activity relationship studies were performed to identify optimal substitution patterns. X-ray structure analysis of a promising dual inhibitor in complex with sEH, as well as molecular docking with LTA(4)H provided a rationale for further optimization. Hereby, scaffold extension was successfully applied to yield potent dual sEH/LTA(4)H inhibitors. The spectrum of pro- and anti-inflammatory lipid mediators was evaluated in M1 and M2 macrophages, stimulated with LPS, and incubated with the most promising compound 14. The effect of 14 on the inflammatory lipid mediator profile characterizes dual sEH/LTA(4)H inhibitors as an interesting option for future anti-inflammatory agent investigations.
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- 2019
11. Computer-Aided Selective Optimization of Side Activities of Talinolol
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Kerstin Hiesinger, Jan S. Kramer, Janosch Achenbach, Daniel Moser, Julia Weber, Sandra K. Wittmann, Christophe Morisseau, Carlo Angioni, Gerd Geisslinger, Astrid S. Kahnt, Astrid Kaiser, Anna Proschak, Dieter Steinhilber, Denys Pogoryelov, Karen Wagner, Bruce D. Hammock, Ewgenij Proschak, and Publica
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polypharmacology ,Pain Research ,Organic Chemistry ,Selective optimization of side activities ,Pharmacology and Pharmaceutical Sciences ,soluble epoxide hydrolase ,Biochemistry ,Medicinal and Biomolecular Chemistry ,5.1 Pharmaceuticals ,Drug Discovery ,structure-based drug design ,Chronic Pain ,Development of treatments and therapeutic interventions ,computer-aided drug design ,Metabolic and endocrine - Abstract
[Image: see text] Selective optimization of side activities is a valuable source of novel lead structures in drug discovery. In this study, a computer-aided approach was used to deorphanize the pleiotropic cholesterol-lowering effects of the beta-blocker talinolol, which result from the inhibition of the enzyme soluble epoxide hydrolase (sEH). X-ray structure analysis of the sEH in complex with talinolol enables a straightforward optimization of inhibitory potency. The resulting lead structure exhibited in vivo activity in a rat model of diabetic neuropatic pain.
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- 2019
12. LptC from Anabaena sp. PCC 7120: Expression, purification and crystallization
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Enrico Schleiff, Giang Ngo, Denys Pogoryelov, Özkan Yildiz, Martin Centola, and Ganna Krasnoselska
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0106 biological sciences ,Gene Expression ,Crystallography, X-Ray ,medicine.disease_cause ,01 natural sciences ,03 medical and health sciences ,Protein Domains ,Affinity chromatography ,010608 biotechnology ,Protein purification ,medicine ,Escherichia coli ,030304 developmental biology ,0303 health sciences ,biology ,Chemistry ,Periplasmic space ,biology.organism_classification ,Anabaena ,Recombinant Proteins ,Transmembrane domain ,Biochemistry ,bacteria ,Periplasmic Proteins ,Proteobacteria ,Bacterial outer membrane ,Bacteria ,Biotechnology - Abstract
Lipopolysaccharides are central elements of the outer leaflet of the outer membrane of Gram-negative bacteria and as such, of cyanobacteria. In the past, the structural analysis of the system in proteobacteria like Escherichia coli has contributed to a deep understanding of the transport of lipopolysaccharides from plasma membrane to the outer membrane. While many components of the transport system are conserved between proteobacteria and cyanobacteria, the periplasmic LptC appears to be distinct. The cyanobacterial proteins are twice as long as the proteobacterial proteins or proteins from firmicutes. This prompted the question whether the structure of the cyanobacterial proteins is comparable the one of the proteobacterial proteins. To address this question, we expressed LptC from Anabaena sp. PCC 7120 in E. coli as truncated protein without the transmembrane segment. We purified the protein utilizing HIS-tag based affinity chromatography and polished the protein after removal of the tag by size exclusion chromatography. The purified recombinant protein was crystallized by the sitting-drop vapor diffusion technique and best crystals, despite being twinned, diffracted to a resolution of 2.6 A.
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- 2020
13. Ribosome biogenesis factor Tsr3 is the aminocarboxypropyl transferase responsible for 18S rRNA hypermodification in yeast and humans
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Jan Philip Wurm, Jens Wöhnert, Peter Kötter, Denis L. J. Lafontaine, Sunny Sharma, Carina Immer, Britta Meyer, Denys Pogoryelov, and Karl-Dieter Entian
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Models, Molecular ,0301 basic medicine ,S-Adenosylmethionine ,Saccharomyces cerevisiae ,Ribosome biogenesis ,Crystallography, X-Ray ,Ribosome ,Pseudouridine ,03 medical and health sciences ,chemistry.chemical_compound ,23S ribosomal RNA ,Catalytic Domain ,RNA, Ribosomal, 18S ,Genetics ,Humans ,Transferase ,RNA Processing, Post-Transcriptional ,Alkyl and Aryl Transferases ,biology ,Nucleic Acid Enzymes ,Inverted Repeat Sequences ,Hydrogen Bonding ,Ribosomal RNA ,HCT116 Cells ,biology.organism_classification ,3. Good health ,030104 developmental biology ,Biochemistry ,chemistry ,Transfer RNA ,Biologie ,Protein Binding - Abstract
The chemically most complex modification in eukaryotic rRNA is the conserved hypermodified nucleotide N1-methyl-N3-aminocarboxypropyl-pseudouridine (m/acp 3 Ψ) located next to the P-site tRNA on the small subunit 18S rRNA. While S-adenosylmethionine was identified as the source of the aminocarboxypropyl (acp) group more than 40 years ago the enzyme catalyzing the acp transfer remained elusive. Here we identify the cytoplasmic ribosome biogenesis protein Tsr3 as the responsible enzyme in yeast and human cells. In functionally impaired Tsr3-mutants, a reduced level of acp modification directly correlates with increased 20S pre-rRNA accumulation. The crystal structure of archaeal Tsr3 homologs revealed the same fold as in SPOUT-class RNA-methyltransferases but a distinct SAM binding mode. This unique SAM binding mode explains why Tsr3 transfers the acp and not the methyl group of SAM to its substrate. Structurally, Tsr3 therefore represents a novel class of acp transferase enzymes., SCOPUS: ar.j, info:eu-repo/semantics/published
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- 2016
14. Discovery of novel inhibitors of the phosphatase activity of the soluble epoxide hydrolase
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Jan S. Kramer, Ewgenij Proschak, Stefan Knapp, Stefano Woltersdorf, Franca Maria Klingler, Daniel Merk, Apirat Chaikuad, Denys Pogoryelov, G. Enrico Rovati, Jan Heering, Kerstin Hiesinger, Dieter Steinhilber, and Sandra K. Wittmann
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0301 basic medicine ,chemistry.chemical_classification ,Epoxide hydrolase 2 ,Stereochemistry ,Phosphatase ,Antiparallel (biochemistry) ,Biochemistry ,03 medical and health sciences ,030104 developmental biology ,Enzyme ,chemistry ,Hydrolase ,cardiovascular system ,Genetics ,Molecular Biology ,Biotechnology - Abstract
The soluble epoxide Hydrolase (sEH) is an emerging pharmacological target. The enzyme is an antiparallel homodimer, where each of the two subunits is composed of a C-terminal hydrolase domain conne...
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- 2018
15. Discovery of polar spirocyclic orally bioavailable urea inhibitors of soluble epoxide hydrolase
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Markus M. Forsberg, Irene Burghardt, Natalia Kalinchenkova, Jarkko Rautio, Denys Pogoryelov, Igor Konstantinov, Lilia Weizel, Ewgenij Proschak, Mikhail Krasavin, Jukka Leppänen, Darya Bagnyukova, Konstantin Falahati, Jan S. Kramer, Nikolay Zhurilo, Seppo Auriola, Jouni Ihalainen, Victor Hernandez-Olmos, Alexey Lukin, Markus Hartmann, Dmitry Dar'in, and Publica
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0301 basic medicine ,Epoxide hydrolase 2 ,Stereochemistry ,Biochemistry ,Inhibitory Concentration 50 ,03 medical and health sciences ,chemistry.chemical_compound ,Drug Discovery ,Hydrolase ,Humans ,Urea ,Spiro Compounds ,Enzyme Inhibitors ,Molecular Biology ,IC50 ,Epoxide Hydrolases ,Chemistry ,Organic Chemistry ,Recombinant Proteins ,Bioavailability ,Molecular Docking Simulation ,030104 developmental biology ,Solubility ,Lipophilicity ,Enantiomer ,Lead compound - Abstract
Spirocyclic 1-oxa-9-azaspiro[5.5]undecan-4-amine scaffold was explored as a basis for the design of potential inhibitors of soluble epoxide hydrolase (sEH). Synthesis and testing of the initial SAR-probing library followed by biochemical testing against sEH allowed nominating a racemic lead compound (±)-22. The latter showed remarkable (> 0.5 mM) solubility in aqueous phosphate buffer solution, unusually low (for sEH inhibitors) lipophilicity as confirmed by experimentally determined logD7.4 of 0.99, and an excellent oral bioavailability in mice (as well as other pharmacokinetic characteristics). Individual enantiomer profiling revealed that the inhibitory potency primarily resided with the dextrorotatory eutomer (+)-22 (IC50 4.99 ± 0.18 nM). For the latter, a crystal structure of its complex with a C-terminal domain of sEH was obtained and resolved. These data fully validate (+)-22 as a new non-racemic advanced lead compound for further development as a potential therapeutic agent for use in such areas as cardiovascular disease, inflammation and pain.
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- 2018
16. Approved Drugs Containing Thiols as Inhibitors of Metallo-β-lactamases: Strategy To Combat Multidrug-Resistant Bacteria
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Müller Hf, Stephan Göttig, Ewgenij Proschak, Cuesta-Bernal J, Denia Frank, Arno Koenigs, Klaas M. Pos, Franca-Maria Klingler, Hanno Sjuts, Jasmin El-Delik, Thomas A. Wichelhaus, and Denys Pogoryelov
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Models, Molecular ,Imipenem ,medicine.drug_class ,Antibiotics ,Antimicrobial susceptibility ,Pharmacology ,Crystallography, X-Ray ,beta-Lactams ,beta-Lactam Resistance ,beta-Lactamases ,Metallo β lactamase ,Drug Resistance, Multiple, Bacterial ,Drug Discovery ,Escherichia coli ,polycyclic compounds ,Screening method ,medicine ,Humans ,Pseudomonas Infections ,Sulfhydryl Compounds ,Escherichia coli Infections ,chemistry.chemical_classification ,biochemical phenomena, metabolism, and nutrition ,bacterial infections and mycoses ,Anti-Bacterial Agents ,Klebsiella Infections ,Klebsiella pneumoniae ,Enzyme ,Multidrug resistant bacteria ,chemistry ,Pseudomonas aeruginosa ,Molecular Medicine ,beta-Lactamase Inhibitors ,medicine.drug - Abstract
Resistance to β-lactam antibiotics can be mediated by metallo-β-lactamase enzymes (MBLs). An MBL inhibitor could restore the effectiveness of β-lactams. We report on the evaluation of approved thiol-containing drugs as inhibitors of NDM-1, VIM-1, and IMP-7. Drugs were assessed by a novel assay using a purchasable fluorescent substrate and thermal shift. Best compounds were tested in antimicrobial susceptibility assay. Using these orthogonal screening methods, we identified drugs that restored the activity of imipenem.
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- 2015
17. PENG: A Neural Gas-Based Approach for Pharmacophore Elucidation. Method Design, Validation, and Virtual Screening for Novel Ligands of LTA4H
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Jan S. Kramer, René Blöcher, Sandra K. Wittmann, Ewgenij Proschak, Daniel Moser, Janosch Achenbach, and Denys Pogoryelov
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Models, Molecular ,Neural gas ,General Chemical Engineering ,Library and Information Sciences ,Crystallography, X-Ray ,Ligands ,Aminopeptidases ,LigandScout ,Leukotriene-A4 hydrolase ,Hydrolase ,Humans ,Prospective Studies ,Enzyme Inhibitors ,Epoxide hydrolase ,Epoxide Hydrolases ,Virtual screening ,Ligand ,Chemistry ,Reproducibility of Results ,General Chemistry ,Combinatorial chemistry ,High-Throughput Screening Assays ,Computer Science Applications ,Benchmarking ,Thermodynamics ,Neural Networks, Computer ,Pharmacophore ,Algorithms ,Protein Binding - Abstract
The pharmacophore concept is commonly employed in virtual screening for hit identification. A pharmacophore is generally defined as the three-dimensional arrangement of the structural and physicochemical features of a compound responsible for its affinity to a pharmacological target. Given a number of active ligands binding to a particular target in the same manner, it can reasonably be assumed that they have some shared features, a common pharmacophore. We present a growing neural gas (GNG)-based approach for the extraction of the relevant features which we called PENG (pharmacophore elucidation by neural gas). Results of retrospective validation indicate an acceptable quality of the generated models. Additionally a prospective virtual screening for leukotriene A4 hydrolase (LTA4H) inhibitors was performed. LTA4H is a bifunctional zinc metalloprotease which displays both epoxide hydrolase and aminopeptidase activity. We could show that the PENG approach is able to predict the binding mode of the ligand by X-ray crystallography. Furthermore, we identified a novel chemotype of LTA4H inhibitors.
- Published
- 2015
18. FGF1‐mediated cardiomyocyte cell cycle reentry depends on the interaction of FGFR‐1 and Fn14
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Tatyana Novoyatleva, Felix B. Engel, Chinmoy Patra, Amna Sajjad, Denys Pogoryelov, and Ralph T. Schermuly
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animal structures ,Biology ,Fibroblast growth factor ,Biochemistry ,Receptors, Tumor Necrosis Factor ,Genetics ,medicine ,Animals ,Myocytes, Cardiac ,Receptor, Fibroblast Growth Factor, Type 1 ,Fibroblast ,Receptor ,Molecular Biology ,Protein kinase B ,PI3K/AKT/mTOR pathway ,Cell Proliferation ,Cell Cycle ,Membrane Proteins ,Cytokine TWEAK ,FGF1 ,Cell cycle ,Rats ,Cell biology ,Fibroblast Growth Factors ,medicine.anatomical_structure ,TWEAK Receptor ,Fibroblast growth factor receptor ,Tumor Necrosis Factors ,embryonic structures ,Fibroblast Growth Factor 1 ,Apoptosis Regulatory Proteins ,Signal Transduction ,Biotechnology - Abstract
Fibroblast growth factors (FGFs) signal through FGF receptors (FGFRs) mediating a broad range of cellular functions during embryonic development, as well as disease and regeneration during adulthood. Thus, it is important to understand the underlying molecular mechanisms that modulate this system. Here, we show that FGFR-1 can interact with the TNF receptor superfamily member fibroblast growth factor-inducible molecule 14 (Fn14) resulting in cardiomyocyte cell cycle reentry. FGF1-induced cell cycle reentry in neonatal cardiomyocytes could be blocked by Fn14 inhibition, while TWEAK-induced cell cycle activation was inhibited by blocking FGFR-1 signaling. In addition, costimulation experiments revealed a synergistic effect of FGF1 and TWEAK in regard to cardiomyocyte cell cycle induction via PI3K/Akt signaling. Overexpression of Fn14 with either FGFR-1 long [FGFR-1(L)] or FGFR-1 short [FGFR-1(S)] isoforms resulted after FGF1/TWEAK stimulation in cell cycle reentry of >40% adult cardiomyocytes. Finally, coimmunoprecipitation and proximity ligation assays indicated that endogenous FGFR-1 and Fn14 interact with each other in cardiomyocytes. This interaction was strongly enhanced in the presence of their corresponding ligands, FGF1 and TWEAK. Taken together, our data suggest that FGFR-1/Fn14 interaction may represent a novel endogenous mechanism to modulate the action of these receptors and their ligands and to control cardiomyocyte cell cycle reentry.
- Published
- 2014
19. BBA70 of Borrelia burgdorferi Is a Novel Plasminogen-binding Protein
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Brian Stevenson, Diana Barthel, Dominik Kugelstadt, Denys Pogoryelov, Claudia Hammerschmidt, Arno Koenigs, Peter F. Zipfel, Peter Kraiczy, Reinhard Wallich, Christine Skerka, and Brandon L. Jutras
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Proteases ,Plasmin ,Plasma protein binding ,Biochemistry ,Kringle domain ,Microbiology ,Bacterial Proteins ,medicine ,Humans ,Fibrinolysin ,Borrelia burgdorferi ,Molecular Biology ,Complement component 5 ,Lyme Disease ,biology ,Complement C5 ,Plasminogen ,Molecular Bases of Disease ,Cell Biology ,bacterial infections and mycoses ,biology.organism_classification ,Urokinase-Type Plasminogen Activator ,Immunity, Innate ,Protein Structure, Tertiary ,Complement system ,Complement C3b ,bacteria ,Plasminogen activator ,Protein Binding ,medicine.drug - Abstract
The Lyme disease spirochete Borrelia burgdorferi lacks endogenous, surface-exposed proteases. In order to efficiently disseminate throughout the host and penetrate tissue barriers, borreliae rely on recruitment of host proteases, such as plasmin(ogen). Here we report the identification of a novel plasminogen-binding protein, BBA70. Binding of plasminogen is dose-dependent and is affected by ionic strength. The BBA70-plasminogen interaction is mediated by lysine residues, primarily located in a putative C-terminal α-helix of BBA70. These lysine residues appear to interact with the lysine-binding sites in plasminogen kringle domain 4 because a deletion mutant of plasminogen lacking that domain was unable to bind to BBA70. Bound to BBA70, plasminogen activated by urokinase-type plasminogen activator was able to degrade both a synthetic chromogenic substrate and the natural substrate fibrinogen. Furthermore, BBA70-bound plasmin was able to degrade the central complement proteins C3b and C5 and inhibited the bacteriolytic effects of complement. Consistent with these functional activities, BBA70 is located on the borrelial outer surface. Additionally, serological evidence demonstrated that BBA70 is produced during mammalian infection. Taken together, recruitment and activation of plasminogen could play a beneficial role in dissemination of B. burgdorferi in the human host and may possibly aid the spirochete in escaping the defense mechanisms of innate immunity.
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- 2013
20. Microscopic rotary mechanism of ion translocation in the Fo complex of ATP synthases
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Alexander Krah, Özkan Yildiz, José D. Faraldo-Gómez, Julian David Langer, Thomas Meier, and Denys Pogoryelov
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Models, Molecular ,Spectrometry, Mass, Electrospray Ionization ,Proton ,Protein Conformation ,Ion ,Membrane Lipids ,Molecular dynamics ,Protein structure ,X-Ray Diffraction ,ATP synthase gamma subunit ,Spirulina ,Lipid bilayer ,Molecular Biology ,Ions ,Binding Sites ,biology ,Chemistry ,Molecular Motor Proteins ,Cell Biology ,Hydrogen-Ion Concentration ,Transmembrane protein ,Proton-Translocating ATPases ,Crystallography ,Dicyclohexylcarbodiimide ,biology.protein ,Thermodynamics ,Chemical stability ,Protons ,Crystallization ,Energy Metabolism - Abstract
The microscopic mechanism of coupled c-ring rotation and ion translocation in F(1)F(o)-ATP synthases is unknown. Here we present conclusive evidence supporting the notion that the ability of c-rings to rotate within the F(o) complex derives from the interplay between the ion-binding sites and their nonhomogenous microenvironment. This evidence rests on three atomic structures of the c(15) rotor from crystals grown at low pH, soaked at high pH and, after N,N'-dicyclohexylcarbodiimide (DCCD) modification, resolved at 1.8, 3.0 and 2.2 Å, respectively. Alongside a quantitative DCCD-labeling assay and free-energy molecular dynamics calculations, these data demonstrate how the thermodynamic stability of the so-called proton-locked state is maximized by the lipid membrane. By contrast, a hydrophilic environment at the a-subunit-c-ring interface appears to unlock the binding-site conformation and promotes proton exchange with the surrounding solution. Rotation thus occurs as c-subunits stochastically alternate between these environments, directionally biased by the electrochemical transmembrane gradient.
- Published
- 2010
21. Structural and energetic basis for H+ versus Na+ binding selectivity in ATP synthase Fo rotors
- Author
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Julian David Langer, Denys Pogoryelov, José D. Faraldo-Gómez, Thomas Meier, Peter J. Bond, and Alexander Krah
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Models, Molecular ,Spectrometry, Mass, Electrospray Ionization ,Protein Conformation ,Stereochemistry ,Biophysics ,Biochemistry ,Fusobacteria ,Proton-motive force ,03 medical and health sciences ,Ion binding ,Protein structure ,c-subunit ring rotor ,Ion selectivity ,Molecular dynamics simulation ,Spirulina ,Binding site ,Electrochemical gradient ,F1Fo ATP synthase ,Binding selectivity ,Dicyclohexylcarbodiimide modification ,030304 developmental biology ,Fo rotor ,0303 health sciences ,Binding Sites ,Sodium-motive force ,ATP synthase ,biology ,Mass spectrometry ,Chemiosmosis ,Chemistry ,030302 biochemistry & molecular biology ,Cell Biology ,Membrane protein structure ,Amino Acid Substitution ,Bacterial Proton-Translocating ATPases ,Mutagenesis, Site-Directed ,biology.protein ,Membrane bioenergetics ,Free-energy calculation ,Thermodynamics ,Mutant Proteins ,Selectivity - Abstract
The functional mechanism of the F1Fo ATP synthase, like many membrane transporters and pumps, entails a conformational cycle that is coupled to the movement of H+ or Na+ ions across its transmembrane domain, down an electrochemical gradient. This coupling is an efficient means of energy transduction and regulation, provided that ion binding to the membrane domain, known as Fo, is appropriately selective. In this study we set out to establish the structural and energetic basis for the ion-binding selectivity of the membrane-embedded Fo rotors of two representative ATP synthases. First, we use a biochemical approach to demonstrate the inherent binding selectivity of these rotors, that is, independently from the rest of the enzyme. We then use atomically detailed computer simulations of wild-type and mutagenized rotors to calculate and rationalize their selectivity, on the basis of the structure, dynamics and coordination chemistry of the binding sites. We conclude that H+ selectivity is most likely a robust property of all Fo rotors, arising from the prominent presence of a conserved carboxylic acid and its intrinsic chemical propensity for protonation, as well as from the structural plasticity of the binding sites. In H+-coupled rotors, the incorporation of hydrophobic side chains to the binding sites enhances this inherent H+ selectivity. Size restriction may also favor H+ over Na+, but increasing size alone does not confer Na+ selectivity. Rather, the degree to which Fo rotors may exhibit Na+ coupling relies on the presence of a sufficient number of suitable coordinating side chains and/or structural water molecules. These ligands accomplish a shift in the relative binding energetics, which under some physiological conditions may be sufficient to provide Na+ dependence.
- Published
- 2010
- Full Text
- View/download PDF
22. High-resolution structure of the rotor ring of a proton-dependent ATP synthase
- Author
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Özkan Yildiz, Thomas Meier, Denys Pogoryelov, and José D. Faraldo-Gómez
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Proton ,Protein subunit ,Molecular Sequence Data ,Static Electricity ,Molecular Conformation ,Protonation ,Protein structure ,Structural Biology ,ATP synthase gamma subunit ,Cations ,Spirulina ,Amino Acid Sequence ,Molecular Biology ,Ions ,Binding Sites ,Sequence Homology, Amino Acid ,biology ,ATP synthase ,Chemistry ,Sodium ,Biological Transport ,Lipids ,Protein Structure, Tertiary ,Crystallography ,Bacterial Proton-Translocating ATPases ,Helix ,biology.protein ,Protons ,ATP synthase alpha/beta subunits - Abstract
The crystal structure of the c-ring from the proton-coupled F1Fo ATP synthase from Spirulina platensis is shown at 2.1-A resolution. The ring includes 15 membrane-embedded c subunits forming an hourglass-shaped assembly. The structure demonstrates that proton translocation across the membrane entails protonation of a conserved glutamate located near the membrane center in the c subunit outer helix. The proton is locked in this site by a precise hydrogen bond network reminiscent of that in Na+-dependent ATP synthases. However, the structure suggests that the different coordination chemistry of the bound proton and the smaller curvature of the outer helix drastically enhance the selectivity of the H+ site against other cations, including H3O+. We propose a model for proton translocation whereby the c subunits remain in this proton-locked state when facing the membrane lipid. Proton exchange would occur in a more hydrophilic and electrostatically distinct environment upon contact with the a subunit interface.
- Published
- 2009
23. Probing the rotor subunit interface of the ATP synthase from Ilyobacter tartaricus
- Author
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Uwe Schlattner, Konstantin Pervushin, Denys Pogoryelov, Peter Dimroth, Yaroslav Nikolaev, and Thomas Meier
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0303 health sciences ,ATP synthase ,biology ,Chemistry ,Protein subunit ,Kinetics ,Sequence alignment ,Cell Biology ,Biochemistry ,03 medical and health sciences ,Crystallography ,0302 clinical medicine ,Docking (molecular) ,biology.protein ,Surface plasmon resonance ,Molecular Biology ,Peptide sequence ,030217 neurology & neurosurgery ,030304 developmental biology ,Gamma subunit - Abstract
The interaction between the c(11)ring and the gammaepsilon complex, forming the rotor of the Ilyobacter tartaricus ATP synthase, was probed by surface plasmon resonance spectroscopy and in vitro reconstitution analysis. The results provide, for the first time, a direct and quantitative assessment of the stability of the rotor. The data indicated very tight binding between the c(11)ring and the gammaepsilon complex, with an apparent K(d) value of approximately 7.4nm. The rotor assembly was primarily dependent on the interaction of the cring with the gammasubunit, and binding of the cring to the free epsilon subunit was not observed. Mutagenesis of selected conserved amino acid residues of all three rotor components (cR45, cQ46, gammaE204, gammaF203 and epsilonH38) severely affected rotor assembly. The interaction kinetics between the gammaepsilon complex and c(11)ring mutants suggested that the assembly of the c(11)gammaepsiloncomplex was governed by interactions of low and high affinity. Low-affinity binding was observed between the polar loops of the cring subunits and the bottom part of the gamma subunit. High-affinity interactions, involving the two residues gammaE204 and epsilonH38, stabilized the holo-c(11)gammaepsilon complex. NMR experiments indicated the acquisition of conformational order in otherwise flexible C- and N-terminal regions of the gamma subunit on rotor assembly. The results of this study suggest that docking of the central stalk of the F(1)complex to the rotor ring of F(o) to form tight, but reversible, contacts provides an explanation for the relative ease of dissociation and reconstitution of F(1)F(o)complexes.
- Published
- 2008
24. A tridecameric c ring of the adenosine triphosphate (ATP) synthase from the thermoalkaliphilic Bacillus sp. strain TA2.A1 facilitates ATP synthesis at low electrochemical proton potential
- Author
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Gregory M. Cook, Thomas Meier, Doreen Matthies, Peter Dimroth, Nina Morgner, Bernhard Brutschy, Stefanie Keis, and Denys Pogoryelov
- Subjects
chemistry.chemical_classification ,biology ,ATP synthase ,Strain (chemistry) ,Chemiosmosis ,Stereochemistry ,Protein subunit ,Proton-Motive Force ,Bacillus ,Microbiology ,Protein Subunits ,Proton-Translocating ATPases ,chemistry.chemical_compound ,Adenosine Triphosphate ,Enzyme ,Bacterial Proteins ,chemistry ,Biochemistry ,biology.protein ,Protons ,Molecular Biology ,Adenosine triphosphate ,ATP synthase alpha/beta subunits ,Electrochemical potential - Abstract
Despite the thermodynamic problem imposed on alkaliphilic bacteria of synthesizing adenosine triphosphate (ATP) against a large inverted pH gradient and consequently a low electrochemical proton potential, these bacteria still utilize a proton-coupled F(1)F(o)-ATP synthase to synthesize ATP. One potential solution to this apparent thermodynamic problem would be the operation of a larger oligomeric c ring, which would raise the ion to ATP ratio, thus facilitating the conversion of a low electrochemical potential into a significant phosphorylation potential. To address this hypothesis, we have purified the oligomeric c ring from the thermoalkaliphilic bacterium Bacillus sp. strain TA2.A1 and determined the number of c-subunits using a novel mass spectrometry method, termed 'laser-induced liquid bead ion desorption' (LILBID). This technique allows the mass determination of non-covalently assembled, detergent-solubilized membrane protein complexes, and hence enables an accurate determination of c ring stoichiometries. We show that the Bacillus sp. strain TA2.A1 ATP synthase harbours a tridecameric c ring. The operation of a c ring with 13 subunits renders the thermodynamic problem of ATP synthesis at alkaline pH less severe and may represent a strategy for ATP synthesis at low electrochemical potential.
- Published
- 2007
25. The Oligomeric State of c Rings from Cyanobacterial F-ATP Synthases Varies from 13 to 15
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Adriana L. Klyszejko, Christian Reichen, Daniel J. Müller, Peter Dimroth, René A. Brunisholz, Thomas Meier, Denys Pogoryelov, University of Zurich, and Dimroth, P
- Subjects
Cyanobacteria ,Stereochemistry ,610 Medicine & health ,10071 Functional Genomics Center Zurich ,Microbiology ,Oligomer ,chemistry.chemical_compound ,1312 Molecular Biology ,Molecular Biology ,Chroococcales ,Gel electrophoresis ,Nostocales ,biology ,ATP synthase ,2404 Microbiology ,biology.organism_classification ,Enzymes and Proteins ,Protein Subunits ,Proton-Translocating ATPases ,chemistry ,Biochemistry ,biology.protein ,570 Life sciences ,Oscillatoriales ,U7 Systems Biology / Functional Genomics ,Stoichiometry - Abstract
We isolated the c rings of F-ATP synthases from eight cyanobacterial strains belonging to four different taxonomic classes ( Chroococcales , Nostocales , Oscillatoriales , and Gloeobacteria ). These c rings showed different mobilities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), probably reflecting their molecular masses. This supposition was validated with the previously characterized c 11 , c 14 , and c 15 rings, which migrated on SDS-PAGE in proportion to their molecular masses. Hence, the masses of the cyanobacterial c rings can conveniently be deduced from their electrophoretic mobilities and, together with the masses of the c monomers, allow the calculation of the c ring stoichiometries. The method is a simple and fast way to determine stoichiometries of SDS-stable c rings and hence a convenient means to unambiguously determine the ion-to-ATP ratio, a parameter reflecting the bioenergetic efficacy of F-ATP synthases. AFM imaging was used to prove the accuracy of the method and confirmed that the c ring of Synechococcus elongatus SAG 89.79 is a tridecameric oligomer. Despite the high conservation of the c-subunit sequences from cyanobacterial strains from various environmental groups, the stoichiometries of their c rings varied between c 13 and c 15 . This systematic study of the c-ring stoichiometries suggests that variability of c-ring sizes might represent an adaptation of the individual cyanobacterial species to their particular environmental and physiological conditions. Furthermore, the two new examples of c 15 rings underline once more that an F 1 /F o symmetry mismatch is not an obligatory feature of all F-ATP synthases.
- Published
- 2007
26. On the principle of ion selectivity in Na + /H + -coupled membrane proteins: Experimental and theoretical studies of an ATP synthase rotor
- Author
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Denys Pogoryelov, José D. Faraldo-Gómez, Vanessa Leone, and Thomas Meier
- Subjects
Binding Sites ,Multidisciplinary ,Sequence Homology, Amino Acid ,ATP synthase ,biology ,Protein Conformation ,Chemistry ,Stereochemistry ,Molecular Sequence Data ,Sodium ,Membrane Proteins ,Isothermal titration calorimetry ,Transmembrane protein ,Proton-Translocating ATPases ,Protein structure ,Membrane ,PNAS Plus ,Membrane protein ,biology.protein ,Amino Acid Sequence ,Protons ,Selectivity ,Ion transporter ,Protein Binding - Abstract
Numerous membrane transporters and enzymes couple their mechanisms to the permeation of Na(+) or H(+), thereby harnessing the energy stored in the form of transmembrane electrochemical potential gradients to sustain their activities. The molecular and environmental factors that control and modulate the ion specificity of most of these systems are, however, poorly understood. Here, we use isothermal titration calorimetry to determine the Na(+)/H(+) selectivity of the ion-driven membrane rotor of an F-type ATP synthase. Consistent with earlier theoretical predictions, we find that this rotor is significantly H(+) selective, although not sufficiently to be functionally coupled to H(+), owing to the large excess of Na(+) in physiological settings. The functional Na(+) specificity of this ATP synthase thus results from two opposing factors, namely its inherent chemical selectivity and the relative availability of the coupling ion. Further theoretical studies of this membrane rotor, and of two others with a much stronger and a slightly weaker H(+) selectivity, indicate that, although the inherent selectivity of their ion-binding sites is largely set by the balance of polar and hydrophobic groups flanking a conserved carboxylic side chain, subtle variations in their structure and conformational dynamics, for a similar chemical makeup, can also have a significant contribution. We propose that the principle of ion selectivity outlined here may provide a rationale for the differentiation of Na(+)- and H(+)-coupled systems in other families of membrane transporters and enzymes.
- Published
- 2015
27. The Effects of Salt Stress on Photosynthetic Electron Transport and Thylakoid Membrane Proteins in the Cyanobacterium Spirulina platensis
- Author
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Gyözö Garab, Denys Pogoryelov, Sistla D.S. Murthy, László Kovács, and Putty-Reddy Sudhir
- Subjects
Photosynthetic reaction centre ,Photoinhibition ,Photosystem II ,Photosystem II Protein Complex ,food and beverages ,Light-harvesting complexes of green plants ,macromolecular substances ,General Medicine ,Sodium Chloride ,Biology ,Cyanobacteria ,Photosystem I ,Thylakoids ,Biochemistry ,Electron Transport ,Thylakoid ,Light-dependent reactions ,Photosynthesis ,Molecular Biology ,Chlorophyll fluorescence - Abstract
The response of Spirulina (Arthrospira) platensis to high salt stress was investigated by incubating the cells in light of moderate intensity in the presence of 0.8 M NaCl. NaCl caused a decrease in photosystem II (PSII) mediated oxygen evolution activity and increase in photosystem I (PSI) activity and the amount of P700. Similarly maximal efficiency of PSII (Fv/Fm) and variable fluorescence (Fv/Fo) were also declined in salt-stressed cells. Western blot analysis reveal that the inhibition in PSII activity is due to a 40 % loss of a thylakoid membrane protein, known as D1, which is located in PSII reaction center. NaCl treatment of cells also resulted in the alterations of other thylakoid membrane proteins: most prominently, a dramatic diminishment of the 47-kDa chlorophyll protein (CP) and 94-kDa protein, and accumulation of a 17-kDa protein band were observed in SDS-PAGE. The changes in 47-kDa and 94-kDa proteins lead to the decreased energy transfer from light harvesting antenna to PSII, which was accompanied by alterations in the chlorophyll fluorescence emission spectra of whole cells and isolated thylakoids. Therefore we conclude that salt stress has various effects on photosynthetic electron transport activities due to the marked alterations in the composition of thylakoid membrane proteins.
- Published
- 2005
28. [Untitled]
- Author
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Zoltán Gombos, P.-R. Sudhir, László Kovács, I. Brown, Győző Garab, and Denys Pogoryelov
- Subjects
Photosystem II ,biology ,Physiology ,Antiporter ,Sodium ,Intracellular pH ,chemistry.chemical_element ,Cell Biology ,biology.organism_classification ,Photosynthesis ,Redox ,Electron transport chain ,chemistry ,Biochemistry ,Biophysics ,Arthrospira - Abstract
Arthrospira (Spirulina) platensis (A. platensis) is a model organism for investigation of adaptation of photosynthetic organisms to extreme environmental conditions: the cell functions in this cyanobacterium are optimized to high pH and high concentration (150–250 mM) of Na+. However, the mechanism of the possible fine-tuning of the photosynthetic functions to these extreme conditions and/or the regulation of the cellular environment to optimize the photosynthetic functions is poorly understood. In this work we investigated the effect of Na-ions on different photosynthetic activities: linear electron transport reactions (measured by means of polarography and spectrophotometry), the activity of photosystem II (PS II) (thermoluminescence and chlorophyll a fluorescence induction), and redox turnover of the cytochrome b6f complex (flash photolysis); and measured the changes of the intracellular pH (9-aminoacridine fluorescence). It was found that sodium deprivation of cells in the dark at pH 10 inhibited, within 40 min, all measured photosynthetic reactions, and led to an alkalinization of the intracellular pH, which rose from the physiological value of about 8.3–9.6. These were partially and totally restored by readdition of Na-ions at 2.5–25 mM and about 200 mM, respectively. The intracellular pH and the photosynthetic functions were also sensitive to monensin, an exogenous Na+/H+ exchanger, which collapses both proton and sodium gradients across the cytoplasmic membrane. These observations explain the strict Na+-dependency of the photosynthetic electron transport at high extracellular pH, provide experimental evidence on the alkalization of the intracellular environment, and support the hypothesized role of an Na+/H+ antiport through the plasma membrane in pH homeostasis (Schlesinger et al. (1996). J. Phycol.32, 608–613). Further, we show that (i) the specific site of inactivation of the photosynthetic electron transport at alkaline pH is to be found at the water splitting enzyme; (ii) in contrast to earlier reports, the inactivation occurs in the dark and, for short periods, without detectable damage in the photosynthetic apparatus; and (iii) in contrast to high pH, Na+ dependency in the neutral pH range is shown not to originate from PSII, but from the acceptor side of PSI. These data permit us to conclude that the intracellular environment rather than the machinery of the photosynthetic electron transport is adjusted to the extreme conditions of high pH and high Na+ concentration.
- Published
- 2003
29. Experimental Determination of the Ion Selectivity of an ATP-Synthase Membrane Rotor by Isothermal Titration Calorimetry
- Author
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José D. Faraldo-Gómez, Vanessa Leone, Ernst Grell, Thomas Meier, and Denys Pogoryelov
- Subjects
biology ,ATP synthase ,Sodium ,ATPase ,Analytical chemistry ,Biophysics ,chemistry.chemical_element ,Isothermal titration calorimetry ,Crystallography ,Membrane ,chemistry ,biology.protein ,Titration ,Electrochemical gradient ,Selectivity - Abstract
ATP synthases are rotating nanomachines that couple ATP synthesis or hydrolysis to the transmembrane flow of protons or sodium ions down or against their electrochemical gradient. The key coupling element is a membrane-embedded subcomplex, the c-ring. We have recently proposed a principle that explains the ion selectivity of the c-ring, and validated this through functional studies of several ATP synthases. Specifically, we have proposed that a conserved Glu/Asp confers a universal H+ selectivity to all c-ring binding sites, and that additional amino-acids, which vary among species, have evolved to modulate this selectivity. In particular, we have shown that polar groups can suppress the H+ selectivity of the c-ring by a factor of 1-103. Thus, the enzyme becomes coupled to Na+, due to the large excess of Na+ over H+ under physiological conditions. Here, we further assess this theory by directly measuring the selectivity of a representative c-ring through Isothermal Titration Calorimetry. Specifically, we characterized the c-ring from the ATP synthase of Ilyobacter tartaricus. From titrations at different pH values, we established that Kd(Na+) ∼ 0.3 mM while Kd(H+) ∼ 0.3 μM, confirming the notion that this prototypical Na+-coupled c-ring is in fact H+ selective, although to a much smaller degree than those actually H+-driven. Comparing our results with those obtained for Enterococcus hirae demonstrates that, as predicted by our theory, the I. tartaricus c-ring is 100 times less Na+ selective. The weaker affinity for Na+ of the I.tartaricus ring is also coherent with the 10-fold difference in Km(Na+) values between these enzymes, at high pH. Taken together, these experiments demonstrate that the c-ring is the main determinant of the physiological ion specificity of rotary ATPases, and provide a conclusive validation of our theory.
- Published
- 2014
- Full Text
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30. ATP Synthase Structure
- Author
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Thomas Meier and Denys Pogoryelov
- Published
- 2013
31. Tuning the ion specificity of the ATP synthase rotor
- Author
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Ganna O. Krasnoselska, Thomas Meier, José D. Faraldo-Gómez, Alexander Krah, Denys Pogoryelov, and Julian David Langer
- Subjects
ATP synthase ,biology ,Rotor (electric) ,law ,Chemistry ,biology.protein ,Biophysics ,Cell Biology ,Biochemistry ,Ion ,law.invention - Published
- 2012
- Full Text
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32. Structural basis for the ion selectivity of F-ATP-synthase c-ring rotors
- Author
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Denys Pogoryelov, José D. Faraldo-Gómez, Thomas Meier, Alexander Krah, Peter J. Bond, and Julian David Langer
- Subjects
Ion selectivity ,ATP synthase ,biology ,Stereochemistry ,Chemistry ,biology.protein ,Biophysics ,Cell Biology ,Biochemistry ,ATP synthase alpha/beta subunits - Published
- 2010
- Full Text
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33. Complete Ion-Coordination Structure in the Rotor Ring of Na+ -Dependent F-ATP Synthases
- Author
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José D. Faraldo-Gómez, Denys Pogoryelov, Kay Diederichs, Alexander Krah, Peter J. Bond, and Thomas Meier
- Subjects
Binding Sites ,Chemistry ,Stereochemistry ,Protein Conformation ,Sodium ,Protein Data Bank (RCSB PDB) ,FiFo-ATP synthase rotor ,Ligand (biochemistry) ,Ring (chemistry) ,Crystallography, X-Ray ,Fusobacteria ,llyobacter tartaricus ,Crystallography ,Molecular dynamics ,Structural Biology ,Bacterial Proton-Translocating ATPases ,ddc:570 ,sodium-motive force ,Molecule ,Binding site ,Selectivity ,Lipid bilayer ,ion coordination and selectivity ,Molecular Biology ,coring structure - Abstract
Summary The membrane-embedded rotors of Na + -dependent F-ATP synthases comprise 11 c-subunits that form a ring, with 11 Na + binding sites in between adjacent subunits. Following an updated crystallographic analysis of the c-ring from Ilyobacter tartaricus , we report the complete ion-coordination structure of the Na + sites. In addition to the four residues previously identified, there exists a fifth ligand, namely, a buried structural water molecule. This water is itself coordinated by Thr67, which, sequence analysis reveals, is the only residue involved in binding that distinguishes Na + synthases from H + -ATP synthases known to date. Molecular dynamics simulations and free-energy calculations of the c-ring in a lipid membrane lend clear support to the notion that this fifth ligand is a water molecule, and illustrate its influence on the selectivity of the binding sites. Given the evolutionary ascendancy of sodium over proton bioenergetics, this structure uncovers an ancient strategy for selective ion coupling in ATP synthases.
- Published
- 2009
34. S1.35 The rotor subunit interface of the ATP synthase from Ilyobacter tartaricus
- Author
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Peter Dimroth, Denys Pogoryelov, Uwe Schlattner, and Thomas Meier
- Subjects
biology ,ATP synthase ,Rotor (electric) ,Chemistry ,Protein subunit ,Ilyobacter tartaricus ,Biophysics ,Cell Biology ,Biochemistry ,law.invention ,ATP synthase gamma subunit ,law ,biology.protein - Published
- 2008
- Full Text
- View/download PDF
35. Correction to Approved Drugs Containing Thiols as Inhibitors of Metallo-β-lactamases: Strategy To Combat Multidrug-Resistant Bacteria
- Author
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Jenifer Cuesta-Bernal, Arno Koenigs, Stephan Göttig, H Florian Müller, Denys Pogoryelov, Ewgenij Proschak, Franca-M Klingler, Hanno Sjuts, Denia Frank, Thomas A. Wichelhaus, Jasmin El-Delik, and Klaas M. Pos
- Subjects
Multidrug resistant bacteria ,Chemistry ,Drug Discovery ,Molecular Medicine ,Bioinformatics ,Metallo β lactamase ,Microbiology - Published
- 2015
36. The c15 ring of the Spirulina platensis F-ATP synthase: F1/F0 symmetry mismatch is not obligatory
- Author
-
Thomas Meier, Peter Dimroth, Janet Vonck, Jinshu Yu, Denys Pogoryelov, and Daniel J. Müller
- Subjects
Spirulina (genus) ,Cyanobacteria ,biology ,ATP synthase ,Stereochemistry ,Chemistry ,Protein subunit ,Scientific Report ,biology.organism_classification ,Mass spectrometry ,Ring (chemistry) ,Microscopy, Atomic Force ,Biochemistry ,Mass Spectrometry ,chemistry.chemical_compound ,Protein Subunits ,Proton-Translocating ATPases ,Monomer ,Genetics ,biology.protein ,Carboxylate ,Crystallization ,Molecular Biology - Abstract
The oligomeric c ring of the F-ATP synthase from the alkaliphilic cyanobacterium Spirulina platensis was isolated and characterized. Mass spectroscopy analysis indicated a mass of 8,210 Da, reflecting that of a c monomer. The mass increased by 206 Da after treatment with the c-subunit-specific inhibitor dicyclohexylcarbodiimide (DCCD), which indicated modification of the ion-binding carboxylate by DCCD. Atomic force microscopy topographs of c rings from S. platensis showed 15 symmetrically assembled subunits. The c15-mer reported here is the largest c ring that is isolated and does not show the classical c-ring mismatch to the three-fold symmetry of the F1 domain.
- Published
- 2005
37. S1.44 Mass determination of membrane protein complexes in detergent solution: The c rings from F-ATP synthases
- Author
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Nina Morgner, Thomas Meier, Doreen Matthies, Denys Pogoryelov, and Bernd Brutschy
- Subjects
Membrane protein ,Biochemistry ,biology ,Chemiosmosis ,Chemistry ,ATP synthase gamma subunit ,biology.protein ,Biophysics ,Cell Biology ,Integral membrane protein - Full Text
- View/download PDF
38. S1.34 What determines the size of the Fo rotor?
- Author
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Thomas Meier and Denys Pogoryelov
- Subjects
Physics ,Rotor (electric) ,law ,Control theory ,Biophysics ,Cell Biology ,Biochemistry ,law.invention - Full Text
- View/download PDF
39. Engineering rotor ring stoichiometries in the ATP synthase
- Author
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Thomas Meier, Eva-Maria Heller, Julian David Langer, José D. Faraldo-Gómez, Janet Vonck, Daniel J. Müller, Ganna O. Krasnoselska, Vanessa Leone, Adriana L. Klyszejko, and Denys Pogoryelov
- Subjects
0106 biological sciences ,Models, Molecular ,Stereochemistry ,Protein Conformation ,Proteolipids ,Biophysics ,010402 general chemistry ,Ring (chemistry) ,Microscopy, Atomic Force ,01 natural sciences ,Biochemistry ,Molecular dynamics ,chemistry.chemical_compound ,03 medical and health sciences ,Protein structure ,Adenosine Triphosphate ,ATP synthase gamma subunit ,030304 developmental biology ,0303 health sciences ,Multidisciplinary ,biology ,ATP synthase ,Chemistry ,Cell Biology ,Surface Plasmon Resonance ,0104 chemical sciences ,ATP Synthetase Complexes ,Microscopy, Electron ,PNAS Plus ,Mutation ,biology.protein ,Electrophoresis, Polyacrylamide Gel ,Adenosine triphosphate ,ATP synthase alpha/beta subunits ,010606 plant biology & botany - Abstract
ATP synthase membrane rotors consist of a ring of c-subunits whose stoichiometry is constant for a given species but variable across different ones. We investigated the importance of c/c-subunit contacts by site-directed mutagenesis of a conserved stretch of glycines (GxGxGxGxG) in a bacterial c 11 ring. Structural and biochemical studies show a direct, specific influence on the c-subunit stoichiometry, revealing c , c 12 , c 13 , c 14 , and c >14 rings. Molecular dynamics simulations rationalize this effect in terms of the energetics and geometry of the c-subunit interfaces. Quantitative data from a spectroscopic interaction study demonstrate that the complex assembly is independent of the c-ring size. Real-time ATP synthesis experiments in proteoliposomes show the mutant enzyme, harboring the larger c 12 instead of c 11 , is functional at lower ion motive force. The high degree of compliance in the architecture of the ATP synthase rotor offers a rationale for the natural diversity of c-ring stoichiometries, which likely reflect adaptations to specific bioenergetic demands. These results provide the basis for bioengineering ATP synthases with customized ion-to-ATP ratios, by sequence modifications.
- Full Text
- View/download PDF
40. Correction to ApprovedDrugs Containing Thiols as Inhibitors of Metallo-β-lactamases:Strategy To Combat Multidrug-Resistant Bacteria.
- Author
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Franca-M. Klingler, ThomasA. Wichelhaus, Denia Frank, Jenifer Cuesta-Bernal, Jasmin El-Delik, H. Florian Müller, Hanno Sjuts, Stephan Göttig, Arno Koenigs, Klaas M. Pos, Denys Pogoryelov, and Ewgenij Proschak
- Published
- 2015
- Full Text
- View/download PDF
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