Long-Range RNA Structural Information via a Paramagnetically Tagged Reporter Protein
| Autor: | Nico Tjandra, Rohith Rajasekaran, Jan Marchant, Guillermo A. Bermejo, Madeleine Strickland, Colin T. O’Hern, Michael F. Summers, Jonathan Catazaro, Marie-Paule Strub |
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| Rok vydání: | 2019 |
| Předmět: |
Models
Molecular RNA metabolism chemistry.chemical_classification Base Sequence Chemistry Magnetic Phenomena RNA General Chemistry 010402 general chemistry 01 natural sciences Biochemistry Article Catalysis In vitro Insert (molecular biology) Ribonucleoprotein U1 Small Nuclear 0104 chemical sciences Colloid and Surface Chemistry Biophysics Nucleic Acid Conformation Nucleotide Base sequence Binding site Ribonucleoprotein |
| Zdroj: | Journal of the American Chemical Society. 141:1430-1434 |
| ISSN: | 1520-5126 0002-7863 |
| Popis: | NMR has provided a wealth of structural and dynamical information for RNA molecules of up to ∼50 nucleotides, but its application to larger RNAs has been hampered in part by difficulties establishing global structural features. A potential solution involves measurement of NMR perturbations after site-specific paramagnetic labeling. Although the approach works well for proteins, the inability to place the label at specific sites has prevented its application to larger RNAs transcribed in vitro. Here, we present a strategy in which RNA loop residues are modified to promote binding to a paramagnetically tagged reporter protein. Lanthanide-induced pseudocontact shifts are demonstrated for a 232-nucleotide RNA bound to tagged derivatives of the spliceosomal U1A RNA-binding domain. Further, the method is validated with a 36-nucleotide RNA for which measured NMR values agreed with predictions based on the previously known protein and RNA structures. The ability to readily insert U1A binding sites into ubiquitous hairpin and/or loop structures should make this approach broadly applicable for the atomic-level study of large RNAs. |
| Databáze: | OpenAIRE |
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