EPR; Single-molecule spectroscopy; Intrinsically disordered regions; Cell biology; Phase separation; NMR; RNA-binding proteins; Molecular pathogenesis
Zosel Franziska, Soranno Andrea, Buholzer Karin J., Nettels Daniel, Schuler Benjamin (2020), Depletion interactions modulate the binding between disordered proteins in crowded environments, in
Proceedings of the National Academy of Sciences, 117(24), 13480-13489.
Zosel Franziska, Mercadante Davide, Nettels Daniel, Schuler Benjamin (2018), A proline switch explains kinetic heterogeneity in a coupled folding and binding reaction, in
Nature Communications, 9(1), 3332-3332.
Grotz Kara K., Nueesch Mark F., Holmstrom Erik D., Heinz Marcel, Stelzl Lukas S., Schuler Benjamin, Hummer Gerhard (2018), Dispersion Correction Alleviates Dye Stacking of Single-Stranded DNA and RNA in Simulations of Single-Molecule Fluorescence Experiments, in
The Journal of Physical Chemistry B, 122(49), 11626-11639.
Holmstrom Erik D., Nettels Daniel, Schuler Benjamin (2018), Conformational Plasticity of Hepatitis C Virus Core Protein Enables RNA-Induced Formation of Nucleocapsid-like Particles, in
Journal of Molecular Biology, 430(16), 2453-2467.
Holmstrom Erik D., Holla Andrea, Zheng Wenwei, Nettels Daniel, Best Robert B., Schuler Benjamin (2018), Accurate Transfer Efficiencies, Distance Distributions, and Ensembles of Unfolded and Intrinsically Disordered Proteins From Single-Molecule FRET, in Cruzer Zoe (ed.), Elsevier, USA, 287-325.
Zosel Franziska, Haenni Dominik, Soranno Andrea, Nettels Daniel, Schuler Benjamin (2017), Combining short- and long-range fluorescence reporters with simulations to explore the intramolecular dynamics of an intrinsically disordered protein, in
The Journal of Chemical Physics, 147(15), 152708-152708.
Plitzko Jürgen M, Schuler Benjamin, Selenko Philipp (2017), Structural Biology outside the box — inside the cell, in
Current Opinion in Structural Biology, 46, 110-121.
ZoselFranziska, HollaAndrea, SchulerBenjamin,
Labeling of proteins for single-molecule fluorescence spectroscopy, Springer, Ney York.
| Author |
Masliah, Gregoire |
| Publication date |
21.02.2018 |
| Persistent Identifier (PID) |
https://www.rcsb.org/structure/5N8M |
| Repository |
RCSB Protein Data Bank
|
| Abstract |
The accurate cleavage of pre-micro(mi)RNAs by Dicer and mi/siRNA guide strand selection are important steps in forming the RNA-induced silencing complex (RISC). The role of Dicer binding partner TRBP in these processes remains poorly understood. Here, we solved the solution structure of the two N-terminal dsRNA binding domains (dsRBDs) of TRBP in complex with a functionally asymmetric siRNA using NMR, EPR, and single-molecule spectroscopy. We find that siRNA recognition by the dsRBDs is not sequence-specific but rather depends on the RNA shape. The two dsRBDs can swap their binding sites, giving rise to two equally populated, pseudo-symmetrical complexes, showing that TRBP is not a primary sensor of siRNA asymmetry. Using our structure to model a Dicer-TRBP-siRNA ternary complex, we show that TRBP's dsRBDs and Dicer's RNase III domains bind a canonical 19 base pair siRNA on opposite sides, supporting a mechanism whereby TRBP influences Dicer-mediated cleavage accuracy by binding the dsRNA region of the pre-miRNA during Dicer cleavage.
| Author |
Masliah, Gregoire |
| Publication date |
21.02.2018 |
| Persistent Identifier (PID) |
https://www.rcsb.org/structure/5N8L |
| Repository |
RCSB Protein Data Bank
|
| Abstract |
The accurate cleavage of pre-micro(mi)RNAs by Dicer and mi/siRNA guide strand selection are important steps in forming the RNA-induced silencing complex (RISC). The role of Dicer binding partner TRBP in these processes remains poorly understood. Here, we solved the solution structure of the two N-terminal dsRNA binding domains (dsRBDs) of TRBP in complex with a functionally asymmetric siRNA using NMR, EPR, and single-molecule spectroscopy. We find that siRNA recognition by the dsRBDs is not sequence-specific but rather depends on the RNA shape. The two dsRBDs can swap their binding sites, giving rise to two equally populated, pseudo-symmetrical complexes, showing that TRBP is not a primary sensor of siRNA asymmetry. Using our structure to model a Dicer-TRBP-siRNA ternary complex, we show that TRBP's dsRBDs and Dicer's RNase III domains bind a canonical 19 base pair siRNA on opposite sides, supporting a mechanism whereby TRBP influences Dicer-mediated cleavage accuracy by binding the dsRNA region of the pre-miRNA during Dicer cleavage.
A surprisingly large fraction of proteins contain extended regions or domains that lack a stable three-dimensional structure under physiological conditions. It is becoming increasingly clear that these intrinsically disordered regions (IDRs) play important roles in many cellular functions and are correspondingly often involved in pathological processes. A particularly important and large group of molecules with high abundance of IDRs are proteins involved in RNA binding and processing. However, the functional relevance of these conserved regions and their impact on disease development are largely unclear, to a large part because of the experimental challenges in investigating systems with such large conformational heterogeneity and pronounced dynamics. In this collaborative project, we combine the expertise of four prominent research groups with highly complementary expertise, both conceptually and methodologically, to address this elusive issue. We bring together the most important methods for the investigation of proteins containing large unstructured regions and their interactions and functional properties, both in vitro and in vivo: nuclear magnetic resonance spectroscopy (NMR), single-molecule Förster resonance energy transfer (FRET), electron paramagnetic resonance spectroscopy (EPR), biochemistry methods, human cells, organotypic slice cultures, and fluorescence microscopy. Conceptually, our groups integrate leading expertise in the function and conformational properties of RNA, RNA-binding proteins, conformationally heterogeneous systems, and the role of RNA-binding proteins in the molecular pathogenesis of neurodegeneration.In this unique combination, we will •establish the methodology for obtaining a complete picture of the structural ensembles of RNA-binding proteins containing disordered regions,•elucidate the effect of intrinsically disordered regions on protein-RNA and protein-protein interactions and their functional consequences,•identify the role and structural properties of IDRs for protein-RNA interactions involved in liquid-liquid phase separation and stress granule formation,•characterize how the transition from liquid-like droplets to pathogenic inclusions is controlled by protein-RNA interactions and intrinsically disordered regions.