Abstract - Vormann

Novel anti-viral agents are needed in response to the severe acute respiratory syndrome coronavirus 2 (Sars-CoV-2) pandemic. One potential drug target is the multi-domain nonstructural protein 3 (nsp3). Nsp3 is involved in forming the Sars-Cov-2 replication organelle (RO), which drives the coronavirus replication in the host cell. Nsp3 is one major constituent of molecular pores embedded in the Double Membrane Vesicles (DMVs) bilayer. These DMV pores allow the import and export of viral RNA. As the largest protein encoded by the coronavirus, nsp3 comprises 16 domains, including the N-terminal Ubiquitin-like domain 1 (Ubl1), forming the cytosolic-faced prongs of the crown-like pore. Considering that ubiquitin-like domains are involved in the regulation of diverse biological processes, the list of potential viral and host cell interaction partners is numerous. Currently, information on the structure, dynamics, and binding partners of nsp3 is limited. Thus, our in vitro studies aim for structural and dynamic information on nsp3-Ubl1 by nuclear magnetic resonance (NMR) spectroscopy and other biophysical methods. For this purpose, we have expressed and purified the 15N, 13C isotopically labeled Ubl1 domain and characterized its internal and conformational dynamics by NMR spectroscopy. Additionally, circular dichroism (CD) spectroscopy was performed to analyze the secondary structural elements of Ubl1.

Katharina Vormann 1,2, Tobias Stief 1,2, Ralf Bartenschlager 3, Nils-Alexander Lakomek 1,2 1 Forschungszentrum Jülich, Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Jülich, Germany 2 Heinrich Heine University Düsseldorf, Institute of Physical Biology, Düsseldorf, Germany 3 Deutsches Krebsforschungszentrum (DKFZ), Division Virus-Associated Carcinogenesis, Heidelberg, Germany