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2. 1H, 13C and 15N chemical shift assignment of the stem-loops 5b + c from the 5′-UTR of SARS-CoV-2

Author

Mertinkus, Klara R., Grün, J. Tassilo, Altincekic, Nadide, Bains, Jasleen Kaur, Ceylan, Betül, Ferner, Jan-Peter, Frydman, Lucio, Fürtig, Boris, Hengesbach, Martin, Hohmann, Katharina F., Hymon, Daniel, Kim, Jihyun, Knezic, Božana, Novakovic, Mihajlo, Oxenfarth, Andreas, Peter, Stephen A., Qureshi, Nusrat S., Richter, Christian, Scherf, Tali, Schlundt, Andreas, Schnieders, Robbin, Schwalbe, Harald, Stirnal, Elke, Sudakov, Alexey, Vögele, Jennifer, Wacker, Anna, Weigand, Julia E., Wirmer-Bartoschek, Julia, Martin, Maria A. Wirtz, and Wöhnert, Jens

Published
2022
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5. Structure of an internal loop motif with three consecutive U•U mismatches from stem–loop 1 in the 3′-UTR of the SARS-CoV-2 genomic RNA

Author

Vögele, Jennifer, primary, Duchardt-Ferner, Elke, additional, Bains, Jasleen Kaur, additional, Knezic, Bozana, additional, Wacker, Anna, additional, Sich, Christian, additional, Weigand, Julia E, additional, Šponer, Jiří, additional, Schwalbe, Harald, additional, Krepl, Miroslav, additional, and Wöhnert, Jens, additional

Published
2024
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7. Dissecting the Conformational Heterogeneity of Stem-Loop Substructures of the Fifth Element in the 5′-Untranslated Region of SARS-CoV-2

Author

Mertinkus, Klara R., Oxenfarth, Andreas, Richter, Christian, Wacker, Anna, Mata, Carlos P., Carazo, Jose Maria, Schlundt, Andreas, and Schwalbe, Harald

Abstract

Throughout the family of coronaviruses, structured RNA elements within the 5′ region of the genome are highly conserved. The fifth stem-loop element from SARS-CoV-2 (5_SL5) represents an example of an RNA structural element, repeatedly occurring in coronaviruses. It contains a conserved, repetitive fold within its substructures SL5a and SL5b. We herein report the detailed characterization of the structure and dynamics of elements SL5a and SL5b that are located immediately upstream of the SARS-CoV-2 ORF1a/b start codon. Exploiting the unique ability of solution NMR methods, we show that the structures of both apical loops are modulated by structural differences in the remote parts located in their stem regions. We further integrated our high-resolution models of SL5a/b into the context of full-length 5_SL5 structures by combining different structural biology methods. Finally, we evaluated the impact of the two most common VoC mutations within 5_SL5 with respect to individual base-pair stability.

Published
2024
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8. 1H, 13C, 15N and 31P chemical shift assignment for stem-loop 4 from the 5′-UTR of SARS-CoV-2

Author

Vögele, Jennifer, Ferner, Jan-Peter, Altincekic, Nadide, Bains, Jasleen Kaur, Ceylan, Betül, Fürtig, Boris, Grün, J. Tassilo, Hengesbach, Martin, Hohmann, Katharina F., Hymon, Daniel, Knezic, Bozana, Löhr, Frank, Peter, Stephen A., Pyper, Dennis, Qureshi, Nusrat S., Richter, Christian, Schlundt, Andreas, Schwalbe, Harald, Stirnal, Elke, Sudakov, Alexey, Wacker, Anna, Weigand, Julia E., Wirmer-Bartoschek, Julia, Wöhnert, Jens, and Duchardt-Ferner, Elke

Published
2021
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9. 1H, 13C and 15N assignment of stem-loop SL1 from the 5'-UTR of SARS-CoV-2

Author

Richter, Christian, Hohmann, Katharina F., Toews, Sabrina, Mathieu, Daniel, Altincekic, Nadide, Bains, Jasleen Kaur, Binas, Oliver, Ceylan, Betül, Duchardt-Ferner, Elke, Ferner, Jan, Fürtig, Boris, Grün, J. Tassilo, Hengesbach, Martin, Hymon, Daniel, Jonker, Hendrik R. A., Knezic, Bozana, Korn, Sophie M., Landgraf, Tom, Löhr, Frank, Peter, Stephen A., Pyper, Dennis J., Qureshi, Nusrat S., Schlundt, Andreas, Schnieders, Robbin, Stirnal, Elke, Sudakov, Alexey, Vögele, Jennifer, Weigand, Julia E., Wirmer-Bartoschek, Julia, Witt, Kerstin, Wöhnert, Jens, Schwalbe, Harald, and Wacker, Anna

Published
2021
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11. 1H, 13C and 15N chemical shift assignment of the stem-loop 5a from the 5′-UTR of SARS-CoV-2

Author

Schnieders, Robbin, Peter, Stephen A., Banijamali, Elnaz, Riad, Magdalena, Altincekic, Nadide, Bains, Jasleen Kaur, Ceylan, Betül, Fürtig, Boris, Grün, J. Tassilo, Hengesbach, Martin, Hohmann, Katharina F., Hymon, Daniel, Knezic, Bozana, Oxenfarth, Andreas, Petzold, Katja, Qureshi, Nusrat S., Richter, Christian, Schlagnitweit, Judith, Schlundt, Andreas, Schwalbe, Harald, Stirnal, Elke, Sudakov, Alexey, Vögele, Jennifer, Wacker, Anna, Weigand, Julia E., Wirmer-Bartoschek, Julia, and Wöhnert, Jens

Published
2021
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12. Comprehensive Fragment Screening of the SARS‐CoV‐2 Proteome Explores Novel Chemical Space for Drug Development

Author

Berg, Hannes, Wirtz Martin, Maria A., Altincekic, Nadide, Alshamleh, Islam, Kaur Bains, Jasleen, Blechar, Julius, Ceylan, Betül, Jesus, Vanessa de, Dhamotharan, Karthikeyan, Fuks, Christin, Gande, Santosh L., Hargittay, Bruno, Hohmann, Katharina F., Hutchison, Marie T., Korn, Sophie Marianne, Krishnathas, Robin, Kutz, Felicitas, Linhard, Verena, Matzel, Tobias, Meiser, Nathalie, Niesteruk, Anna, Pyper, Dennis J., Schulte, Linda, Trucks, Sven, Azzaoui, Kamal, Blommers, Marcel J. J., Gadiya, Yojana, Karki, Reagon, Zaliani, Andrea, Gribbon, Philip, Silva Almeida, Marcius da, Dinis Anobom, Cristiane, Bula, Anna L., Bütikofer, Matthias, Putinhon Caruso, Ícaro, Caterina Felli, Isabella, Da Poian, Andrea T., Cardoso de Amorim, Gisele, Fourkiotis, Nikolaos K., Gallo, Angelo, Ghosh, Dhiman, Gomes‐Neto, Francisco, Gorbatyuk, Oksana, Hao, Bing, Kurauskas, Vilius, Lecoq, Lauriane, Li, Yunfeng, Cunha Mebus‐Antunes, Nathane, Mompeán, Miguel, Cristtina Neves‐Martins, Thais, Ninot‐Pedrosa, Martí, Pinheiro, Anderson S., Pontoriero, Letizia, Pustovalova, Yulia, Riek, Roland, Robertson, Angus J., Jose Abi Saad, Marie, Treviño, Miguel Á., Tsika, Aikaterini C., Almeida, Fabio C. L., Bax, Ad, Henzler‐Wildman, Katherine, Hoch, Jeffrey C., Jaudzems, Kristaps, Laurents, Douglas V., Orts, Julien, Pierattelli, Roberta, Spyroulias, Georgios A., Duchardt‐Ferner, Elke, Ferner, Jan, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia E., Wirmer‐Bartoschek, Julia, Wöhnert, Jens, Schwalbe, Harald, Berg, Hannes, Wirtz Martin, Maria A., Altincekic, Nadide, Alshamleh, Islam, Kaur Bains, Jasleen, Blechar, Julius, Ceylan, Betül, Jesus, Vanessa de, Dhamotharan, Karthikeyan, Fuks, Christin, Gande, Santosh L., Hargittay, Bruno, Hohmann, Katharina F., Hutchison, Marie T., Korn, Sophie Marianne, Krishnathas, Robin, Kutz, Felicitas, Linhard, Verena, Matzel, Tobias, Meiser, Nathalie, Niesteruk, Anna, Pyper, Dennis J., Schulte, Linda, Trucks, Sven, Azzaoui, Kamal, Blommers, Marcel J. J., Gadiya, Yojana, Karki, Reagon, Zaliani, Andrea, Gribbon, Philip, Silva Almeida, Marcius da, Dinis Anobom, Cristiane, Bula, Anna L., Bütikofer, Matthias, Putinhon Caruso, Ícaro, Caterina Felli, Isabella, Da Poian, Andrea T., Cardoso de Amorim, Gisele, Fourkiotis, Nikolaos K., Gallo, Angelo, Ghosh, Dhiman, Gomes‐Neto, Francisco, Gorbatyuk, Oksana, Hao, Bing, Kurauskas, Vilius, Lecoq, Lauriane, Li, Yunfeng, Cunha Mebus‐Antunes, Nathane, Mompeán, Miguel, Cristtina Neves‐Martins, Thais, Ninot‐Pedrosa, Martí, Pinheiro, Anderson S., Pontoriero, Letizia, Pustovalova, Yulia, Riek, Roland, Robertson, Angus J., Jose Abi Saad, Marie, Treviño, Miguel Á., Tsika, Aikaterini C., Almeida, Fabio C. L., Bax, Ad, Henzler‐Wildman, Katherine, Hoch, Jeffrey C., Jaudzems, Kristaps, Laurents, Douglas V., Orts, Julien, Pierattelli, Roberta, Spyroulias, Georgios A., Duchardt‐Ferner, Elke, Ferner, Jan, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia E., Wirmer‐Bartoschek, Julia, Wöhnert, Jens, and Schwalbe, Harald

Abstract

SARS‐CoV‐2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti‐virals. Within the international Covid19‐NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80 % of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR‐detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure‐based drug design against the SCoV2 proteome.

Published
2024

13. m6A Methylation of Transcription Leader Sequence of SARS‐CoV‐2 Impacts Discontinuous Transcription of Subgenomic mRNAs.

Author

Becker, Matthias A., Meiser, Nathalie, Schmidt‐Dengler, Martina, Richter, Christian, Wacker, Anna, Schwalbe, Harald, and Hengesbach, Martin

Subjects
GENETIC transcription, CYTOSKELETAL proteins, METHYLATION, SARS-CoV-2, VIRAL proteins, MESSENGER RNA
Abstract

The SARS‐CoV‐2 genome has been shown to be m6A methylated at several positions in vivo. Strikingly, a DRACH motif, the recognition motif for adenosine methylation, resides in the core of the transcriptional regulatory leader sequence (TRS−L) at position A74, which is highly conserved and essential for viral discontinuous transcription. Methylation at position A74 correlates with viral pathogenicity. Discontinuous transcription produces a set of subgenomic mRNAs that function as templates for translation of all structural and accessory proteins. A74 is base‐paired in the short stem‐loop structure 5'SL3 that opens during discontinuous transcription to form long‐range RNA‐RNA interactions with nascent (−)‐strand transcripts at complementary TRS‐body sequences. A74 can be methylated by the human METTL3/METTL14 complex in vitro. Here, we investigate its impact on the structural stability of 5'SL3 and the long‐range TRS‐leader:TRS‐body duplex formation necessary for synthesis of subgenomic mRNAs of all four viral structural proteins. Methylation uniformly destabilizes 5'SL3 and long‐range duplexes and alters their relative equilibrium populations, suggesting that the m6A74 modification acts as a regulator for the abundance of viral structural proteins due to this destabilization. [ABSTRACT FROM AUTHOR]

Published
2024
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16. Das COVID19‐NMR‐Konsortium: Ein öffentlicher Bericht über den Einfluss dieser neuen globalen Kollaboration

Author

Duchardt‐Ferner, Elke, primary, Ferner, Jan, additional, Fürtig, Boris, additional, Hengesbach, Martin, additional, Richter, Christian, additional, Schlundt, Andreas, additional, Sreeramulu, Sridhar, additional, Wacker, Anna, additional, Weigand, Julia E., additional, Wirmer‐Bartoschek, Julia, additional, and Schwalbe, Harald, additional

Published
2023
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19. NMR 1H,19F-based screening of the four stem-looped structure 5_SL1–SL4 located in the 5′-untranslated region of SARS-CoV 2 RNAElectronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d3md00322a

Author

Hymon, Daniel, Martins, Jason, Richter, Christian, Sreeramulu, Sridhar, Wacker, Anna, Ferner, Jan, Patwardhan, Neeraj N., Hargrove, Amanda E., and Schwalbe, Harald

Abstract

Development of new antiviral medication against the beta-coronavirus SARS-CoV-2 (SCoV2) is actively being pursued. Both NMR spectroscopy and crystallography as structural screening technologies have been utilised to screen the viral proteome for binding to fragment libraries. Here, we report on NMR screening of elements of the viral RNA genome with two different ligand libraries using 1H-NMR-screening experiments and 1H and 19F NMR-screening experiments for fluorinated compounds. We screened against the 5′-terminal 119 nucleotides located in the 5′-untranslated region of the RNA genome of SCoV2 and further dissected the four stem-loops into its constituent RNA elements to test specificity of binding of ligands to shorter and longer viral RNA stretches. The first library (DRTL-F library) is enriched in ligands binding to RNA motifs, while the second library (DSI-poised library) represents a fragment library originally designed for protein screening. Conducting screens with two different libraries allows us to compare different NMR screening methodologies, describe NMR screening workflows, validate the two different fragment libraries, and derive initial leads for further downstream medicinal chemistry optimisation.

Published
2024
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20. NMR characterization and ligand binding site of the stem–loop 2 motif from the Delta variant of SARS-CoV-2

Author

Matzel, Tobias, Martin, Maria Wirtz, Herr, Alexander, Wacker, Anna, Richter, Christian, Sreeramulu, Sridhar, and Schwalbe, Harald

Abstract

The stem–loop 2 motif (s2m) in SARS-CoV-2 (SCoV-2) is located in the 3′-UTR. Although s2m has been reported to display characteristics of a mobile genomic element that might lead to an evolutionary advantage, its function has remained unknown. The secondary structure of the original SCoV-2 RNA sequence (Wuhan-Hu-1) was determined by NMR in late 2020, delineating the base-pairing pattern and revealing substantial differences in secondary structure compared to SARS-CoV-1 (SCoV-1). The existence of a single G29742–A29756 mismatch in the upper stem of s2m leads to its destabilization and impedes a complete NMR analysis. With Delta, a variant of concern has evolved with one mutation compared to the original sequence that replaces G29742 by U29742. We show here that this mutation results in a more defined structure at ambient temperature accompanied by a rise in melting temperature. Consequently, we were able to identify >90% of the relevant NMR resonances using a combination of selective RNA labeling and filtered 2D NOESY as well as 4D NMR experiments. We present a comprehensive NMR analysis of the secondary structure, (sub)nanosecond dynamics, and ribose conformation of s2m Delta based on heteronuclear 13C NOE and T1measurements and ribose carbon chemical shift-derived canonical coordinates. We further show that the G29742U mutation in Delta has no influence on the druggability of s2m compared to the Wuhan-Hu-1 sequence. With the assignment at hand, we identify the flexible regions of s2m as the primary site for small molecule binding.

Published
2024
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21. Comprehensive Fragment Screening of the SARS‐CoV‐2 Proteome Explores Novel Chemical Space for Drug Development

Author

Berg, Hannes, primary, Wirtz Martin, Maria A., additional, Altincekic, Nadide, additional, Alshamleh, Islam, additional, Kaur Bains, Jasleen, additional, Blechar, Julius, additional, Ceylan, Betül, additional, de Jesus, Vanessa, additional, Dhamotharan, Karthikeyan, additional, Fuks, Christin, additional, Gande, Santosh L., additional, Hargittay, Bruno, additional, Hohmann, Katharina F., additional, Hutchison, Marie T., additional, Marianne Korn, Sophie, additional, Krishnathas, Robin, additional, Kutz, Felicitas, additional, Linhard, Verena, additional, Matzel, Tobias, additional, Meiser, Nathalie, additional, Niesteruk, Anna, additional, Pyper, Dennis J., additional, Schulte, Linda, additional, Trucks, Sven, additional, Azzaoui, Kamal, additional, Blommers, Marcel J. J., additional, Gadiya, Yojana, additional, Karki, Reagon, additional, Zaliani, Andrea, additional, Gribbon, Philip, additional, da Silva Almeida, Marcius, additional, Dinis Anobom, Cristiane, additional, Bula, Anna L., additional, Bütikofer, Matthias, additional, Putinhon Caruso, Ícaro, additional, Caterina Felli, Isabella, additional, Da Poian, Andrea T., additional, Cardoso de Amorim, Gisele, additional, Fourkiotis, Nikolaos K., additional, Gallo, Angelo, additional, Ghosh, Dhiman, additional, Gomes‐Neto, Francisco, additional, Gorbatyuk, Oksana, additional, Hao, Bing, additional, Kurauskas, Vilius, additional, Lecoq, Lauriane, additional, Li, Yunfeng, additional, Cunha Mebus‐Antunes, Nathane, additional, Mompeán, Miguel, additional, Cristtina Neves‐Martins, Thais, additional, Ninot‐Pedrosa, Martí, additional, Pinheiro, Anderson S., additional, Pontoriero, Letizia, additional, Pustovalova, Yulia, additional, Riek, Roland, additional, Robertson, Angus J., additional, Jose Abi Saad, Marie, additional, Treviño, Miguel Á., additional, Tsika, Aikaterini C., additional, Almeida, Fabio C. L., additional, Bax, Ad, additional, Henzler‐Wildman, Katherine, additional, Hoch, Jeffrey C., additional, Jaudzems, Kristaps, additional, Laurents, Douglas V., additional, Orts, Julien, additional, Pierattelli, Roberta, additional, Spyroulias, Georgios A., additional, Duchardt‐Ferner, Elke, additional, Ferner, Jan, additional, Fürtig, Boris, additional, Hengesbach, Martin, additional, Löhr, Frank, additional, Qureshi, Nusrat, additional, Richter, Christian, additional, Saxena, Krishna, additional, Schlundt, Andreas, additional, Sreeramulu, Sridhar, additional, Wacker, Anna, additional, Weigand, Julia E., additional, Wirmer‐Bartoschek, Julia, additional, Wöhnert, Jens, additional, and Schwalbe, Harald, additional

Published
2022
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22. Comprehensive Fragment Screening of the SARS-CoV-2 Proteome Explores Novel Chemical Space for Drug Development

Author

State of Hesse, German Research Foundation, European Commission, Ministero dell'Istruzione, dell'Università e della Ricerca, Agence Nationale de la Recherche (France), Centre National de la Recherche Scientifique (France), National Institutes of Health (US), National Science Foundation (US), Latvian Council of Science, Berg, Hannes [0000-0002-2060-4296], Wirtz Martin, Maria A. [0000-0002-0318-7785], Altincekic, Nadide [0000-0001-6370-3414], Alshamleh, Islam [0000-0001-6714-3602], Dhamotharan, Karthikeyan [0000-0003-0226-7350], Marianne Korn, Sophie [0000-0003-3798-3277], Schulte, Linda [0000-0002-9334-8908], da Silva Almeida, Marcius [0000-0003-4921-8185], Caterina Felli, Isabella [0000-0002-6018-9090], Fourkiotis, Nikolaos K. [0000-0002-5197-4142], Gallo, Angelo [0000-0001-9778-4822], Ninot-Pedrosa, Martí [0000-0003-2851-9990], Pontoriero, Letizia [0000-0002-5586-1305], Treviño, Miguel A. [0000-0002-0738-5973], Tsika, Aikaterini C. [000-0002-3723-0606], Almeida, Fabio C.L. [0000-0001-6046-7006], Bax, Ad [0000-0002-9809-5700], Henzler-Wildman, Katherine [0000-0002-5295-2121], Hoch, Jeffrey C. [0000-0002-9230-2019], Jaudzems, Kristaps [0000-0003-3922-2447], Laurents, D.V. [0000-0002-4187-165X], Ferner, Jan [0000-0002-2009-3203], Hengesbach, Martin [0000-0001-9414-1602], Löhr, Frank [0000-0001-6399-9497], Qureshi, Nusrat [0000-0002-5753-5984], Richter, Christian [0000-0002-5420-2826], Schlundt, Andreas [0000-0003-2254-7560], Weigand, Julia E. [0000-0003-4247-1348], Wirmer-Bartoschek, Julia [0000-0002-0642-1311], Schwalbe, Harald [0000-0001-5693-7909], Berg, Hannes, Wirtz Martin, Maria A., Altincekic, Nadide, Alshamleh, Islam, Kaur Bains, Jasleen, Blechar, Julius, Ceylan, Betül, Jesus, Vanessa de, Dhamotharan, Karthikeyan, Fuks, Christin, Gande, Santosh L., Hargittay, Bruno, Hohmann, Katharina F., Hutchison, Marie T., Marianne Korn, Sophie, Krishnathas, Robin, Kutz, Felicitas, Linhard, Verena, Matzel, Tobias, Meiser, Nathalie, Niesteruk, Anna, Pyper, Dennis J., Schulte, Linda, Trucks, Sven, Azzaoui, Kamal, Blommers, Marcel J.J., Gadiya, Yojana, Karki, Reagon, Zaliani, Andrea, Gribbon, Philip, da Silva Almeida, Marcius, Dinis Anobom, Cristiane, Bula, Anna L., Bütikofer, Matthias, Putinhon Caruso, Ícaro, Caterina Felli, Isabella, Da Poian, Andrea T., Cardoso de Amorim, Gisele, Fourkiotis, Nikolaos K., Gallo, Angelo, Ghosh, Dhiman, Gomes-Neto, Francisco, Gorbatyuk, Oksana, Hao, Bing, Kurauskas, Vilius, Lecoq, Lauriane, Li, Yunfeng, Cunha Mebus-Antunes, Nathane, Mompeán, Miguel, Cristtina Neves-Martins, Thais, Ninot-Pedrosa, Martí, Pinheiro, Anderson S.., Pontoriero, Letizia, Pustovalova, Yulia, Riek, Roland, Robertson, Angus J., Jose Abi Saad, Marie, Treviño, Miguel A., Tsika, Aikaterini C., Almeida, Fabio C.L., Bax, Ad, Henzler-Wildman, Katherine, Hoch, Jeffrey C., Jaudzems, Kristaps, Laurents, Douglas V., Orts, Julien, Pierattelli, Roberta, Spyroulias, Georgios A., Duchardt-Ferner, Elke, Ferner, Jan, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia E., Wirmer-Bartoschek, Julia, Wöhnert, Jens, Schwalbe, Harald, State of Hesse, German Research Foundation, European Commission, Ministero dell'Istruzione, dell'Università e della Ricerca, Agence Nationale de la Recherche (France), Centre National de la Recherche Scientifique (France), National Institutes of Health (US), National Science Foundation (US), Latvian Council of Science, Berg, Hannes [0000-0002-2060-4296], Wirtz Martin, Maria A. [0000-0002-0318-7785], Altincekic, Nadide [0000-0001-6370-3414], Alshamleh, Islam [0000-0001-6714-3602], Dhamotharan, Karthikeyan [0000-0003-0226-7350], Marianne Korn, Sophie [0000-0003-3798-3277], Schulte, Linda [0000-0002-9334-8908], da Silva Almeida, Marcius [0000-0003-4921-8185], Caterina Felli, Isabella [0000-0002-6018-9090], Fourkiotis, Nikolaos K. [0000-0002-5197-4142], Gallo, Angelo [0000-0001-9778-4822], Ninot-Pedrosa, Martí [0000-0003-2851-9990], Pontoriero, Letizia [0000-0002-5586-1305], Treviño, Miguel A. [0000-0002-0738-5973], Tsika, Aikaterini C. [000-0002-3723-0606], Almeida, Fabio C.L. [0000-0001-6046-7006], Bax, Ad [0000-0002-9809-5700], Henzler-Wildman, Katherine [0000-0002-5295-2121], Hoch, Jeffrey C. [0000-0002-9230-2019], Jaudzems, Kristaps [0000-0003-3922-2447], Laurents, D.V. [0000-0002-4187-165X], Ferner, Jan [0000-0002-2009-3203], Hengesbach, Martin [0000-0001-9414-1602], Löhr, Frank [0000-0001-6399-9497], Qureshi, Nusrat [0000-0002-5753-5984], Richter, Christian [0000-0002-5420-2826], Schlundt, Andreas [0000-0003-2254-7560], Weigand, Julia E. [0000-0003-4247-1348], Wirmer-Bartoschek, Julia [0000-0002-0642-1311], Schwalbe, Harald [0000-0001-5693-7909], Berg, Hannes, Wirtz Martin, Maria A., Altincekic, Nadide, Alshamleh, Islam, Kaur Bains, Jasleen, Blechar, Julius, Ceylan, Betül, Jesus, Vanessa de, Dhamotharan, Karthikeyan, Fuks, Christin, Gande, Santosh L., Hargittay, Bruno, Hohmann, Katharina F., Hutchison, Marie T., Marianne Korn, Sophie, Krishnathas, Robin, Kutz, Felicitas, Linhard, Verena, Matzel, Tobias, Meiser, Nathalie, Niesteruk, Anna, Pyper, Dennis J., Schulte, Linda, Trucks, Sven, Azzaoui, Kamal, Blommers, Marcel J.J., Gadiya, Yojana, Karki, Reagon, Zaliani, Andrea, Gribbon, Philip, da Silva Almeida, Marcius, Dinis Anobom, Cristiane, Bula, Anna L., Bütikofer, Matthias, Putinhon Caruso, Ícaro, Caterina Felli, Isabella, Da Poian, Andrea T., Cardoso de Amorim, Gisele, Fourkiotis, Nikolaos K., Gallo, Angelo, Ghosh, Dhiman, Gomes-Neto, Francisco, Gorbatyuk, Oksana, Hao, Bing, Kurauskas, Vilius, Lecoq, Lauriane, Li, Yunfeng, Cunha Mebus-Antunes, Nathane, Mompeán, Miguel, Cristtina Neves-Martins, Thais, Ninot-Pedrosa, Martí, Pinheiro, Anderson S.., Pontoriero, Letizia, Pustovalova, Yulia, Riek, Roland, Robertson, Angus J., Jose Abi Saad, Marie, Treviño, Miguel A., Tsika, Aikaterini C., Almeida, Fabio C.L., Bax, Ad, Henzler-Wildman, Katherine, Hoch, Jeffrey C., Jaudzems, Kristaps, Laurents, Douglas V., Orts, Julien, Pierattelli, Roberta, Spyroulias, Georgios A., Duchardt-Ferner, Elke, Ferner, Jan, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia E., Wirmer-Bartoschek, Julia, Wöhnert, Jens, and Schwalbe, Harald

Abstract

SARS-CoV-2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti-virals. Within the international Covid19-NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80 % of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR-detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure-based drug design against the SCoV2 proteome.

Published
2022

24. Exploring the druggability of conserved RNA regulatory elements in the SARS-CoV-2 genome

Author

Sreeramulu, Sridhar, Richter, Christian, Berg, Hannes, Wirtz Martin, Maria A., Ceylan, Betül, Matzel, Tobias, Adam, Jennifer, Altincekic, Nadide, Azzaoui, Kamal, Bains, Jasleen Kaur, Blommers, Marcel Jules José, Ferner, Jan, Fürtig, Boris, Göbel, Michael, Grün, J. Tassilo, Hengesbach, Martin, Hohmann, Katharina F., Hymon, Daniel, Knezic, Bozana, Martins, Jason N., Mertinkus, Klara R., Niesteruk, Anna, Peter, Stephen A., Pyper, Dennis J., Qureshi, Nusrat, Scheffer, Ute, Schlundt, Andreas, Schnieders, Robbin, Stirnal, Elke, Sudakov, Alexey, Tröster, Alix Friederike, Vögele, Jennifer, Wacker, Anna, Weigand, Julia, Wirmer-Bartoschek, Julia, Wöhnert, Jens, and Schwalbe, Harald

Subjects
ddc:570, ddc:540, ddc:610
Abstract

SARS-CoV-2 contains a positive single-stranded RNA genome of approximately 30 000 nucleotides. Within this genome, 15 RNA elements were identified as conserved between SARS-CoV and SARS-CoV-2. By nuclear magnetic resonance (NMR) spectroscopy, we previously determined that these elements fold independently, in line with data from in vivo and ex-vivo structural probing experiments. These elements contain non-base-paired regions that potentially harbor ligand-binding pockets. Here, we performed an NMR-based screening of a poised fragment library of 768 compounds for binding to these RNAs, employing three different 1H-based 1D NMR binding assays. The screening identified common as well as RNA-element specific hits. The results allow selection of the most promising of the 15 RNA elements as putative drug targets. Based on the identified hits, we derive key functional units and groups in ligands for effective targeting of the RNA of SARS-CoV-2.

Published
2021

25. Exploring the Druggability of Conserved RNA Regulatory Elements in the SARS‐CoV‐2 Genome

Author

Sreeramulu, Sridhar, primary, Richter, Christian, additional, Berg, Hannes, additional, Wirtz Martin, Maria A., additional, Ceylan, Betül, additional, Matzel, Tobias, additional, Adam, Jennifer, additional, Altincekic, Nadide, additional, Azzaoui, Kamal, additional, Bains, Jasleen Kaur, additional, Blommers, Marcel J. J., additional, Ferner, Jan, additional, Fürtig, Boris, additional, Göbel, Michael, additional, Grün, J. Tassilo, additional, Hengesbach, Martin, additional, Hohmann, Katharina F., additional, Hymon, Daniel, additional, Knezic, Bozana, additional, Martins, Jason N., additional, Mertinkus, Klara R., additional, Niesteruk, Anna, additional, Peter, Stephen A., additional, Pyper, Dennis J., additional, Qureshi, Nusrat S., additional, Scheffer, Ute, additional, Schlundt, Andreas, additional, Schnieders, Robbin, additional, Stirnal, Elke, additional, Sudakov, Alexey, additional, Tröster, Alix, additional, Vögele, Jennifer, additional, Wacker, Anna, additional, Weigand, Julia E., additional, Wirmer‐Bartoschek, Julia, additional, Wöhnert, Jens, additional, and Schwalbe, Harald, additional

Published
2021
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28. Correction to ‘Secondary structure determination of conserved SARS-CoV-2 RNA elements by NMR spectroscopy’

Author

Wacker, Anna, primary, Weigand, Julia E, additional, Akabayov, Sabine R, additional, Altincekic, Nadide, additional, Bains, Jasleen Kaur, additional, Banijamali, Elnaz, additional, Binas, Oliver, additional, Castillo-Martinez, Jesus, additional, Cetiner, Erhan, additional, Ceylan, Betül, additional, Chiu, Liang-Yuan, additional, Davila-Calderon, Jesse, additional, Dhamotharan, Karthikeyan, additional, Duchardt-Ferner, Elke, additional, Ferner, Jan, additional, Frydman, Lucio, additional, Fürtig, Boris, additional, Gallego, José, additional, Grün, J Tassilo, additional, Hacker, Carolin, additional, Haddad, Christina, additional, Hähnke, Martin, additional, Hengesbach, Martin, additional, Hiller, Fabian, additional, Hohmann, Katharina F, additional, Hymon, Daniel, additional, de Jesus, Vanessa, additional, Jonker, Henry, additional, Keller, Heiko, additional, Knezic, Bozana, additional, Landgraf, Tom, additional, Löhr, Frank, additional, Luo, Le, additional, Mertinkus, Klara R, additional, Muhs, Christina, additional, Novakovic, Mihajlo, additional, Oxenfarth, Andreas, additional, Palomino-Schätzlein, Martina, additional, Petzold, Katja, additional, Peter, Stephen A, additional, Pyper, Dennis J, additional, Qureshi, Nusrat S, additional, Riad, Magdalena, additional, Richter, Christian, additional, Saxena, Krishna, additional, Schamber, Tatjana, additional, Scherf, Tali, additional, Schlagnitweit, Judith, additional, Schlundt, Andreas, additional, Schnieders, Robbin, additional, Schwalbe, Harald, additional, Simba-Lahuasi, Alvaro, additional, Sreeramulu, Sridhar, additional, Stirnal, Elke, additional, Sudakov, Alexey, additional, Tants, Jan-Niklas, additional, Tolbert, Blanton S, additional, Vögele, Jennifer, additional, Weiß, Lena, additional, Wirmer-Bartoschek, Julia, additional, Wirtz Martin, Maria A, additional, Wöhnert, Jens, additional, and Zetzsche, Heidi, additional

Published
2021
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29. In-cell NMR spectroscopy of functional Riboswitch aptamers in eukaryotic cells

Author

Broft, Patrizia, Dzatko, Simon, Krafcikova, Michaela, Wacker, Anna, Hänsel-Hertsch, Robert, Dötsch, Volker, Trantirek, Lukáš, and Schwalbe, Harald

Subjects
ddc:540, ddc:530, ddc:610
Abstract

We report here the in-cell NMR-spectroscopic observation of the binding of the cognate ligand 2′-deoxyguanosine to the aptamer domain of the bacterial 2′-deoxyguanosine-sensing riboswitch in eukaryotic cells, namely Xenopus laevis oocytes and in human HeLa cells. The riboswitch is sufficiently stable in both cell types to allow for detection of binding of the ligand to the riboswitch. Most importantly, we show that the binding mode established by in vitro characterization of this prokaryotic riboswitch is maintained in eukaryotic cellular environment. Our data also bring important methodological insights: Thus far, in-cell NMR studies on RNA in mammalian cells have been limited to investigations of short (

Published
2020

30. 19F NMR-based fragment screening for 14 different biologically active RNAs and 10 DNA and protein counter-screens

Author

Binas, Oliver, Jesus, Vanessa de, Landgraf, Tom, Völklein, Albrecht Eduard, Martins, Jason, Hymon, Daniel, Bains, Jasleen Kaur, Berg, Hannes, Biedenbänder, Thomas, Fürtig, Boris, Gande, Santosh Lakshmi, Niesteruk, Anna, Oxenfarth, Andreas, Qureshi, Nusrat, Schamber, Tatjana, Schnieders, Robbin, Tröster, Alix Friederike, Wacker, Anna, Wirmer-Bartoschek, Julia, Wirtz Martin, Maria Alexandra, Stirnal, Elke, Azzaoui, Kamal, Richter, Christian, Sreeramulu, Sridhar, Blommers, Marcel Jules José, and Schwalbe, Harald

Subjects
ddc:570
Abstract

We report here the nuclear magnetic resonance 19F screening of 14 RNA targets with different secondary and tertiary structure to systematically assess the druggability of RNAs. Our RNA targets include representative bacterial riboswitches that naturally bind with nanomolar affinity and high specificity to cellular metabolites of low molecular weight. Based on counter-screens against five DNAs and five proteins, we can show that RNA can be specifically targeted. To demonstrate the quality of the initial fragment library that has been designed for easy follow-up chemistry, we further show how to increase binding affinity from an initial fragment hit by chemistry that links the identified fragment to the intercalator acridine. Thus, we achieve low-micromolar binding affinity without losing binding specificity between two different terminator structures.

Published
2020

31. Correction to 'Secondary structure determination of conserved SARS-CoV-2 RNA elements by NMR spectroscopy’

Author

Wacker, Anna, Weigand, Julia, Akabayov, Sabine R, Altınçekiç, Nadide, Bains, Jasleen Kaur, Banijamali, Elnaz, Binas, Oliver, Castillo-Martinez, Jesus, Çetiner, Erhan Can, Ceylan, Betül, Chiu, Liang-Yuan, Davila-Calderon, Jesse, Dhamotharan, Karthikeyan, Duchardt-Ferner, Elke, Ferner, Jan, Frydman, Lucio, Fürtig, Boris, Gallego, José, Grün, J. Tassilo, Hacker, Carolin, Haddad, Christina, Hähnke, Martin Jens, Hengesbach, Martin, Hiller, Fabian, Hohmann, Katharina Felicitas, Hymon, Daniel, Jesus, Vanessa de, Jonker, Henry, Keller, Heiko, Knežić, Božana, Landgraf, Tom, Löhr, Frank, Luo, Le, Mertinkus, Klara Rebecca, Muhs, Christina, Novakovic, Mihajlo, Oxenfarth, Andreas, Palomino-Schätzlein, Martina, Petzold, Katja, Peter, Stephen, Pyper, Dennis Joshua, Qureshi, Nusrat, Riad, Magdalena, Richter, Christian, Saxena, Krishna, Schamber, Tatjana, Scherf, Tali, Schlagnitweit, Judith, Schlundt, Andreas, Schnieders, Robbin, Schwalbe, Harald, Simba-Lahuasi, Alvaro, Sreeramulu, Sridhar, Stirnal, Elke, Sudakov, Alexey, Tants, Jan-Niklas, Tolbert, Blanton S, Vögele, Jennifer, Weiß, Lena, Wirmer-Bartoschek, Julia, Wirtz Martin, Maria Alexandra, Wöhnert, Jens, Zetzsche, Heidi, Wacker, Anna, Weigand, Julia, Akabayov, Sabine R, Altınçekiç, Nadide, Bains, Jasleen Kaur, Banijamali, Elnaz, Binas, Oliver, Castillo-Martinez, Jesus, Çetiner, Erhan Can, Ceylan, Betül, Chiu, Liang-Yuan, Davila-Calderon, Jesse, Dhamotharan, Karthikeyan, Duchardt-Ferner, Elke, Ferner, Jan, Frydman, Lucio, Fürtig, Boris, Gallego, José, Grün, J. Tassilo, Hacker, Carolin, Haddad, Christina, Hähnke, Martin Jens, Hengesbach, Martin, Hiller, Fabian, Hohmann, Katharina Felicitas, Hymon, Daniel, Jesus, Vanessa de, Jonker, Henry, Keller, Heiko, Knežić, Božana, Landgraf, Tom, Löhr, Frank, Luo, Le, Mertinkus, Klara Rebecca, Muhs, Christina, Novakovic, Mihajlo, Oxenfarth, Andreas, Palomino-Schätzlein, Martina, Petzold, Katja, Peter, Stephen, Pyper, Dennis Joshua, Qureshi, Nusrat, Riad, Magdalena, Richter, Christian, Saxena, Krishna, Schamber, Tatjana, Scherf, Tali, Schlagnitweit, Judith, Schlundt, Andreas, Schnieders, Robbin, Schwalbe, Harald, Simba-Lahuasi, Alvaro, Sreeramulu, Sridhar, Stirnal, Elke, Sudakov, Alexey, Tants, Jan-Niklas, Tolbert, Blanton S, Vögele, Jennifer, Weiß, Lena, Wirmer-Bartoschek, Julia, Wirtz Martin, Maria Alexandra, Wöhnert, Jens, and Zetzsche, Heidi

Published
2021

32. No Abuse Potential of Silexan in Healthy Recreational Drug Users: A Randomized Controlled Trial

Author

Seifritz, Erich, Möller, Hans-Jürgen, Volz, Hans-Peter, Müller, Walter E, Hopyan, Talar, Wacker, Anna, Schläfke, Sandra, Kasper, Siegfried, Seifritz, Erich, Möller, Hans-Jürgen, Volz, Hans-Peter, Müller, Walter E, Hopyan, Talar, Wacker, Anna, Schläfke, Sandra, and Kasper, Siegfried

Abstract

BACKGROUND Silexan is a lavender essential oil with established anxiolytic and calming efficacy. Here we asked whether there is a potential for abuse in human patients. METHODS We carried out a phase I abuse liability single-center, double-blind, 5-way crossover study in healthy users of recreational central nervous system depressants. They received single oral doses of 80 mg (therapeutic dose) and 640 mg Silexan, 2 mg and 4 mg lorazepam (active control) and placebo in randomized order, with 4- to 14-day washout periods between treatments. Pharmacodynamic measures included validated visual analogue scales assessing positive, negative, and sedative drug effects and balance of effects; a short form of the Addiction Research Center Inventory; and a drug similarity assessment. The primary outcome measure was the individual maximum value on the drug liking visual analogue scale during 24 hours post-dose. RESULTS Forty participants were randomized and 34 were evaluable for pharmacodynamic outcomes. In intraindividual head-to-head comparisons of the drug liking visual analogue scale maximum value, both doses of Silexan were rated similar to placebo whereas differences were observed between Silexan and lorazepam and between placebo and lorazepam (P < .001). These data were supported by all secondary measures of positive drug effects and of balance of effects. Differences between placebo and both doses of Silexan were always negligible in magnitude. Moreover, Silexan showed no sedative effects and was not perceived to be similar to commonly used drugs that participants had used in the past. CONCLUSIONS Silexan did not exhibit any abuse potential in a standard abuse potential detection screen study and is unlikely to be recreationally abused.

Published
2021

33. Characterization of structure and dynamics of the guanidine-II riboswitch from Escherichia coli by NMR spectroscopy and small-angle X-ray scattering (SAXS)

Author

Schamber, Tatjana, Binas, Oliver, Schlundt, Andreas, Wacker, Anna, Schwalbe, Harald, Schamber, Tatjana, Binas, Oliver, Schlundt, Andreas, Wacker, Anna, and Schwalbe, Harald

Abstract

Riboswitches are regulatory RNA elements that undergo functionally important allosteric conformational switching upon binding of specific ligands. The here investigated guanidine-II riboswitch binds the small cation, guanidinium, and forms a kissing loop-loop interaction between its P1 and P2 hairpins. We investigated the structural changes to support previous studies regarding the binding mechanism. Using NMR spectroscopy, we confirmed the structure as observed in crystal structures and we characterized the kissing loop interaction upon addition of Mg2+ and ligand for the riboswitch aptamer from Escherichia coli. We further investigated closely related mutant constructs providing further insight into functional differences between the two (different) hairpins P1 and P2. Formation of intermolecular interactions were probed by small-angle X-ray scattering (SAXS) and NMR DOSY data. All data are consistent and show the formation of oligomeric states of the riboswitch induced by Mg2+ and ligand binding.

Published
2021

37. Secondary structure determination of conserved SARS-CoV-2 RNA elements by NMR spectroscopy

Author

Wacker, Anna, primary, Weigand, Julia E, additional, Akabayov, Sabine R, additional, Altincekic, Nadide, additional, Bains, Jasleen Kaur, additional, Banijamali, Elnaz, additional, Binas, Oliver, additional, Castillo-Martinez, Jesus, additional, Cetiner, Erhan, additional, Ceylan, Betül, additional, Chiu, Liang-Yuan, additional, Davila-Calderon, Jesse, additional, Dhamotharan, Karthikeyan, additional, Duchardt-Ferner, Elke, additional, Ferner, Jan, additional, Frydman, Lucio, additional, Fürtig, Boris, additional, Gallego, José, additional, Grün, J Tassilo, additional, Hacker, Carolin, additional, Haddad, Christina, additional, Hähnke, Martin, additional, Hengesbach, Martin, additional, Hiller, Fabian, additional, Hohmann, Katharina F, additional, Hymon, Daniel, additional, de Jesus, Vanessa, additional, Jonker, Henry, additional, Keller, Heiko, additional, Knezic, Bozana, additional, Landgraf, Tom, additional, Löhr, Frank, additional, Luo, Le, additional, Mertinkus, Klara R, additional, Muhs, Christina, additional, Novakovic, Mihajlo, additional, Oxenfarth, Andreas, additional, Palomino-Schätzlein, Martina, additional, Petzold, Katja, additional, Peter, Stephen A, additional, Pyper, Dennis J, additional, Qureshi, Nusrat S, additional, Riad, Magdalena, additional, Richter, Christian, additional, Saxena, Krishna, additional, Schamber, Tatjana, additional, Scherf, Tali, additional, Schlagnitweit, Judith, additional, Schlundt, Andreas, additional, Schnieders, Robbin, additional, Schwalbe, Harald, additional, Simba-Lahuasi, Alvaro, additional, Sreeramulu, Sridhar, additional, Stirnal, Elke, additional, Sudakov, Alexey, additional, Tants, Jan-Niklas, additional, Tolbert, Blanton S, additional, Vögele, Jennifer, additional, Weiß, Lena, additional, Wirmer-Bartoschek, Julia, additional, Wirtz Martin, Maria A, additional, Wöhnert, Jens, additional, and Zetzsche, Heidi, additional

Published
2020
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38. 19 F NMR‐Based Fragment Screening for 14 Different Biologically Active RNAs and 10 DNA and Protein Counter‐Screens

Author

Binas, Oliver, primary, Jesus, Vanessa, additional, Landgraf, Tom, additional, Völklein, Albrecht Eduard, additional, Martins, Jason, additional, Hymon, Daniel, additional, Kaur Bains, Jasleen, additional, Berg, Hannes, additional, Biedenbänder, Thomas, additional, Fürtig, Boris, additional, Lakshmi Gande, Santosh, additional, Niesteruk, Anna, additional, Oxenfarth, Andreas, additional, Shahin Qureshi, Nusrat, additional, Schamber, Tatjana, additional, Schnieders, Robbin, additional, Tröster, Alix, additional, Wacker, Anna, additional, Wirmer‐Bartoschek, Julia, additional, Wirtz Martin, Maria Alexandra, additional, Stirnal, Elke, additional, Azzaoui, Kamal, additional, Richter, Christian, additional, Sreeramulu, Sridhar, additional, José Blommers, Marcel Jules, additional, and Schwalbe, Harald, additional

Published
2020
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40. 1H, 13C, and 15N backbone chemical shift assignments of coronavirus-2 non-structural protein Nsp10

Author

Kubatova, Nina, Qureshi, Nusrat, Altınçekiç, Nadide, Abele, Rupert, Bains, Jasleen Kaur, Ceylan, Betül, Ferner, Jan, Fuks, Christin, Hargittay, Bruno, Hutchison, Marie, de Jesus, Vanessa, Kutz, Felicitas, Wirtz Martin, Maria Alexandra, Meiser, Nathalie, Linhard, Verena, Pyper, Dennis Joshua, Trucks, Sven, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Schwalbe, Harald, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia, Wirmer-Bartoschek, Julia, Wöhnert, Jens, Kubatova, Nina, Qureshi, Nusrat, Altınçekiç, Nadide, Abele, Rupert, Bains, Jasleen Kaur, Ceylan, Betül, Ferner, Jan, Fuks, Christin, Hargittay, Bruno, Hutchison, Marie, de Jesus, Vanessa, Kutz, Felicitas, Wirtz Martin, Maria Alexandra, Meiser, Nathalie, Linhard, Verena, Pyper, Dennis Joshua, Trucks, Sven, Fürtig, Boris, Hengesbach, Martin, Löhr, Frank, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Schwalbe, Harald, Sreeramulu, Sridhar, Wacker, Anna, Weigand, Julia, Wirmer-Bartoschek, Julia, and Wöhnert, Jens

Abstract

The international Covid19-NMR consortium aims at the comprehensive spectroscopic characterization of SARS-CoV-2 RNA elements and proteins and will provide NMR chemical shift assignments of the molecular components of this virus. The SARS-CoV-2 genome encodes approximately 30 different proteins. Four of these proteins are involved in forming the viral envelope or in the packaging of the RNA genome and are therefore called structural proteins. The other proteins fulfill a variety of functions during the viral life cycle and comprise the so-called non-structural proteins (nsps). Here, we report the near-complete NMR resonance assignment for the backbone chemical shifts of the non-structural protein 10 (nsp10). Nsp10 is part of the viral replication-transcription complex (RTC). It aids in synthesizing and modifying the genomic and subgenomic RNAs. Via its interaction with nsp14, it ensures transcriptional fidelity of the RNA-dependent RNA polymerase, and through its stimulation of the methyltransferase activity of nsp16, it aids in synthesizing the RNA cap structures which protect the viral RNAs from being recognized by the innate immune system. Both of these functions can be potentially targeted by drugs. Our data will aid in performing additional NMR-based characterizations, and provide a basis for the identification of possible small molecule ligands interfering with nsp10 exerting its essential role in viral replication.

Published
2020

41. 1H, 13C, and 15N backbone chemical shift assignments of the apo and the ADP-ribose bound forms of the macrodomain of SARS-CoV-2 non-structural protein 3b

Author

Cantini, F., Banci, L., Altincekic, Nadide, Bains, Jasleen Kaur, Dhamotharan, Karthikeyan, Fuks, Christin, Fürtig, Boris, Gande, S. L., Hargittay, Bruno, Hengesbach, Martin, Hutchison, Marie, Korn, Sophie M., Kubatova, Nina, Kutz, Felicitas, Linhard, Verena, Löhr, Frank, Meiser, Nathalie, Pyper, Dennis Joshua, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Schwalbe, Harald, Sreeramulu, Sridhar, Tants, Jan-Niklas, Wacker, Anna, Weigand, Julia E., Wöhnert, Jens, Tsika, A. C., Fourkiotis, N. K., Spyroulias, G. A., Cantini, F., Banci, L., Altincekic, Nadide, Bains, Jasleen Kaur, Dhamotharan, Karthikeyan, Fuks, Christin, Fürtig, Boris, Gande, S. L., Hargittay, Bruno, Hengesbach, Martin, Hutchison, Marie, Korn, Sophie M., Kubatova, Nina, Kutz, Felicitas, Linhard, Verena, Löhr, Frank, Meiser, Nathalie, Pyper, Dennis Joshua, Qureshi, Nusrat, Richter, Christian, Saxena, Krishna, Schlundt, Andreas, Schwalbe, Harald, Sreeramulu, Sridhar, Tants, Jan-Niklas, Wacker, Anna, Weigand, Julia E., Wöhnert, Jens, Tsika, A. C., Fourkiotis, N. K., and Spyroulias, G. A.

Abstract

The SARS-CoV-2 genome encodes for approximately 30 proteins. Within the international project COVID19-NMR, we distribute the spectroscopic analysis of the viral proteins and RNA. Here, we report NMR chemical shift assignments for the protein Nsp3b, a domain of Nsp3. The 217-kDa large Nsp3 protein contains multiple structurally independent, yet functionally related domains including the viral papain-like protease and Nsp3b, a macrodomain (MD). In general, the MDs of SARS-CoV and MERS-CoV were suggested to play a key role in viral replication by modulating the immune response of the host. The MDs are structurally conserved. They most likely remove ADP-ribose, a common posttranslational modification, from protein side chains. This de-ADP ribosylating function has potentially evolved to protect the virus from the anti-viral ADP-ribosylation catalyzed by poly-ADP-ribose polymerases (PARPs), which in turn are triggered by pathogen-associated sensing of the host immune system. This renders the SARS-CoV-2 Nsp3b a highly relevant drug target in the viral replication process. We here report the near-complete NMR backbone resonance assignment (1H, 13C, 15N) of the putative Nsp3b MD in its apo form and in complex with ADP-ribose. Furthermore, we derive the secondary structure of Nsp3b in solution. In addition, 15N-relaxation data suggest an ordered, rigid core of the MD structure. These data will provide a basis for NMR investigations targeted at obtaining small-molecule inhibitors interfering with the catalytic activity of Nsp3b.

Published
2020

42. Secondary structure determination of conserved SARS-CoV-2 RNA elements by NMR spectroscopy

Author

Wacker, Anna, Weigand, Julia, Akabayov, Sabine R., Altınçekiç, Nadide, Bains, Jasleen Kaur, Banijamali, Elnaz, Binas, Oliver, Castillo-Martinez, Jesus, Çetiner, Erhan Can, Ceylan, Betül, Chiu, Liang-Yuan, Davila-Calderon, Jesse, Dhamotharan, Karthikeyan, Duchardt-Ferner, Elke, Ferner, Jan, Frydman, Lucio, Fürtig, Boris, Gallego, José, Grün, J. Tassilo, Hacker, Carolin, Haddad, Christina, Hähnke, Martin Jens, Hengesbach, Martin, Hiller, Fabian, Hohmann, Katharina Felicitas, Hymon, Daniel, de Jesus, Vanessa, Jonker, Henry, Keller, Heiko, Knežić, Božana, Landgraf, Tom, Löhr, Frank, Luo, Le, Mertinkus, Klara Rebecca, Muhs, Christina, Novakovic, Mihajlo, Oxenfarth, Andreas, Palomino-Schätzlein, Martina, Petzold, Katja, Peter, Stephen, Pyper, Dennis Joshua, Qureshi, Nusrat, Riad, Magdalena, Richter, Christian, Saxena, Krishna, Schamber, Tatjana, Scherf, Tali, Schlagnitweit, Judith, Schlundt, Andreas, Schnieders, Robbin, Schwalbe, Harald, Simba-Lahuasi, Alvaro, Sreeramulu, Sridhar, Stirnal, Elke, Sudakov, Alexey, Tants, Jan-Niklas, Tolbert, Blanton S., Vögele, Jennifer, Weiß, Lena, Wirmer-Bartoschek, Julia, Wirtz Martin, Maria Alexandra, Wöhnert, Jens, Zetzsche, Heidi, Wacker, Anna, Weigand, Julia, Akabayov, Sabine R., Altınçekiç, Nadide, Bains, Jasleen Kaur, Banijamali, Elnaz, Binas, Oliver, Castillo-Martinez, Jesus, Çetiner, Erhan Can, Ceylan, Betül, Chiu, Liang-Yuan, Davila-Calderon, Jesse, Dhamotharan, Karthikeyan, Duchardt-Ferner, Elke, Ferner, Jan, Frydman, Lucio, Fürtig, Boris, Gallego, José, Grün, J. Tassilo, Hacker, Carolin, Haddad, Christina, Hähnke, Martin Jens, Hengesbach, Martin, Hiller, Fabian, Hohmann, Katharina Felicitas, Hymon, Daniel, de Jesus, Vanessa, Jonker, Henry, Keller, Heiko, Knežić, Božana, Landgraf, Tom, Löhr, Frank, Luo, Le, Mertinkus, Klara Rebecca, Muhs, Christina, Novakovic, Mihajlo, Oxenfarth, Andreas, Palomino-Schätzlein, Martina, Petzold, Katja, Peter, Stephen, Pyper, Dennis Joshua, Qureshi, Nusrat, Riad, Magdalena, Richter, Christian, Saxena, Krishna, Schamber, Tatjana, Scherf, Tali, Schlagnitweit, Judith, Schlundt, Andreas, Schnieders, Robbin, Schwalbe, Harald, Simba-Lahuasi, Alvaro, Sreeramulu, Sridhar, Stirnal, Elke, Sudakov, Alexey, Tants, Jan-Niklas, Tolbert, Blanton S., Vögele, Jennifer, Weiß, Lena, Wirmer-Bartoschek, Julia, Wirtz Martin, Maria Alexandra, Wöhnert, Jens, and Zetzsche, Heidi

Abstract

The current pandemic situation caused by the Betacoronavirus SARS-CoV-2 (SCoV2) highlights the need for coordinated research to combat COVID-19. A particularly important aspect is the development of medication. In addition to viral proteins, structured RNA elements represent a potent alternative as drug targets. The search for drugs that target RNA requires their high-resolution structural characterization. Using nuclear magnetic resonance (NMR) spectroscopy, a worldwide consortium of NMR researchers aims to characterize potential RNA drug targets of SCoV2. Here, we report the characterization of 15 conserved RNA elements located at the 5′ end, the ribosomal frameshift segment and the 3′-untranslated region (3′-UTR) of the SCoV2 genome, their large-scale production and NMR-based secondary structure determination. The NMR data are corroborated with secondary structure probing by DMS footprinting experiments. The close agreement of NMR secondary structure determination of isolated RNA elements with DMS footprinting and NMR performed on larger RNA regions shows that the secondary structure elements fold independently. The NMR data reported here provide the basis for NMR investigations of RNA function, RNA interactions with viral and host proteins and screening campaigns to identify potential RNA binders for pharmaceutical intervention.

Published
2020

43. Die Hamburger HafenCity - Leben (in) der Zukunft?

Author

Wacker, Anna Leena, Lange, Jonna, Bandemer, Stella, Wacker, Anna Leena, Lange, Jonna, and Bandemer, Stella

Abstract

Der Hamburger Hafen prägt die Wirtschaft und Optik der Stadt. Der stattfindende Strukturwandel von Gewerbebetrieben zu einer Mischung aus Wohnen und modernen Dienstleistungen soll Image prägend nicht nur für das Gebiet um den Hafen, sondern ganz Hamburg werden. Das städtebauliche Großprojekt HafenCity und insbesondere die Elbphilharmonie werden hier einer Analyse aus kulturanthropologischer Perspektive unterzogen.

Published
2019

44. 19F NMR‐Based Fragment Screening for 14 Different Biologically Active RNAs and 10 DNA and Protein Counter‐Screens.

Author

Binas, Oliver, Jesus, Vanessa, Landgraf, Tom, Völklein, Albrecht Eduard, Martins, Jason, Hymon, Daniel, Kaur Bains, Jasleen, Berg, Hannes, Biedenbänder, Thomas, Fürtig, Boris, Lakshmi Gande, Santosh, Niesteruk, Anna, Oxenfarth, Andreas, Shahin Qureshi, Nusrat, Schamber, Tatjana, Schnieders, Robbin, Tröster, Alix, Wacker, Anna, Wirmer‐Bartoschek, Julia, and Wirtz Martin, Maria Alexandra

Published
2021
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46. Targeting RNA structure in SMN2 reverses spinal muscular atrophy molecular phenotypes

Author

Garcia-Lopez, Amparo, primary, Tessaro, Francesca, additional, Jonker, Hendrik R. A., additional, Wacker, Anna, additional, Richter, Christian, additional, Comte, Arnaud, additional, Berntenis, Nikolaos, additional, Schmucki, Roland, additional, Hatje, Klas, additional, Petermann, Olivier, additional, Chiriano, Gianpaolo, additional, Perozzo, Remo, additional, Sciarra, Daniel, additional, Konieczny, Piotr, additional, Faustino, Ignacio, additional, Fournet, Guy, additional, Orozco, Modesto, additional, Artero, Ruben, additional, Metzger, Friedrich, additional, Ebeling, Martin, additional, Goekjian, Peter, additional, Joseph, Benoît, additional, Schwalbe, Harald, additional, and Scapozza, Leonardo, additional

Published
2018
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48. Targeting RNA structure in SMN2 reverses spinal muscular atrophy molecular phenotypes

Author

Garcia-Lopez, Amparo, Tessaro, Francesca, Jonker, Hendrik R. A., Wacker, Anna, Richter, Christian, Comte, Arnaud, Berntenis, Nikolaos, Schmucki, Roland, Hatje, Klas, Petermann, Olivier, Chiriano, Gianpaolo, Perozzo, Remo, Sciarra, Daniel, Konieczny, Piotr, Faustino, Ignacio, Fournet, Guy, Orozco, Modesto, Artero, Ruben, Metzger, Friedrich, Ebeling, Martin, Goekjian, Peter, Joseph, Benoît, Schwalbe, Harald, Scapozza, Leonardo, Garcia-Lopez, Amparo, Tessaro, Francesca, Jonker, Hendrik R. A., Wacker, Anna, Richter, Christian, Comte, Arnaud, Berntenis, Nikolaos, Schmucki, Roland, Hatje, Klas, Petermann, Olivier, Chiriano, Gianpaolo, Perozzo, Remo, Sciarra, Daniel, Konieczny, Piotr, Faustino, Ignacio, Fournet, Guy, Orozco, Modesto, Artero, Ruben, Metzger, Friedrich, Ebeling, Martin, Goekjian, Peter, Joseph, Benoît, Schwalbe, Harald, and Scapozza, Leonardo

Abstract

Modification of SMN2 exon 7 (E7) splicing is a validated therapeutic strategy against spinal muscular atrophy (SMA). However, a target-based approach to identify small-molecule E7 splicing modifiers has not been attempted, which could reveal novel therapies with improved mechanistic insight. Here, we chose as a target the stem-loop RNA structure TSL2, which overlaps with the 5′ splicing site of E7. A small-molecule TSL2-binding compound, homocarbonyltopsentin (PK4C9), was identified that increases E7 splicing to therapeutic levels and rescues downstream molecular alterations in SMA cells. High-resolution NMR combined with molecular modelling revealed that PK4C9 binds to pentaloop conformations of TSL2 and promotes a shift to triloop conformations that display enhanced E7 splicing. Collectively, our study validates TSL2 as a target for small-molecule drug discovery in SMA, identifies a novel mechanism of action for an E7 splicing modifier, and sets a precedent for other splicing-mediated diseases where RNA structure could be similarly targeted.

Published
2018

49. Pausing guides RNA folding to populate transiently stable RNA structures for riboswitch-based transcription regulation

Author

Steinert, Hannah, primary, Sochor, Florian, additional, Wacker, Anna, additional, Buck, Janina, additional, Helmling, Christina, additional, Hiller, Fabian, additional, Keyhani, Sara, additional, Noeske, Jonas, additional, Grimm, Steffen, additional, Rudolph, Martin M, additional, Keller, Heiko, additional, Mooney, Rachel Anne, additional, Landick, Robert, additional, Suess, Beatrix, additional, Fürtig, Boris, additional, Wöhnert, Jens, additional, and Schwalbe, Harald, additional

Published
2017
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50. Author response: Pausing guides RNA folding to populate transiently stable RNA structures for riboswitch-based transcription regulation

Author

Steinert, Hannah, primary, Sochor, Florian, additional, Wacker, Anna, additional, Buck, Janina, additional, Helmling, Christina, additional, Hiller, Fabian, additional, Keyhani, Sara, additional, Noeske, Jonas, additional, Grimm, Steffen, additional, Rudolph, Martin M, additional, Keller, Heiko, additional, Mooney, Rachel Anne, additional, Landick, Robert, additional, Suess, Beatrix, additional, Fürtig, Boris, additional, Wöhnert, Jens, additional, and Schwalbe, Harald, additional

Published
2017
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