Direct Mapping of Higher-Order RNA Interactions by SHAPE-JuMP
| Autor: | Greggory M. Rice, Kevin M. Weeks, Gillian Houlihan, Matthew J. Smola, Catherine A. Giannetti, Philipp Holliger, Thomas W. Christy, Alain Laederach, Jing Wang, Nikolay V. Dokholyan |
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| Rok vydání: | 2021 |
| Předmět: |
chemistry.chemical_classification
0303 health sciences DNA Complementary Nucleic acid tertiary structure Acylation 030302 biochemistry & molecular biology RNA RNA-Directed DNA Polymerase Sequence Analysis DNA Computational biology Biochemistry Small molecule Article Reverse transcriptase Primer extension 03 medical and health sciences Cross-Linking Reagents chemistry Complementary DNA Oxazines Nucleic Acid Conformation Nucleotide Nucleic acid structure |
| Zdroj: | Biochemistry |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/acs.biochem.1c00270 |
| Popis: | Higher-order structure governs function for many RNAs. However, discerning this structure for large RNA molecules in solution is an unresolved challenge. Here, we present SHAPE-JuMP (selective 2'-hydroxyl acylation analyzed by primer extension and juxtaposed merged pairs) to interrogate through-space RNA tertiary interactions. A bifunctional small molecule is used to chemically link proximal nucleotides in an RNA structure. The RNA cross-link site is then encoded into complementary DNA (cDNA) in a single, direct step using an engineered reverse transcriptase that "jumps" across cross-linked nucleotides. The resulting cDNAs contain a deletion relative to the native RNA sequence, which can be detected by sequencing, that indicates the sites of cross-linked nucleotides. SHAPE-JuMP measures RNA tertiary structure proximity concisely across large RNA molecules at nanometer resolution. SHAPE-JuMP is especially effective at measuring interactions in multihelix junctions and loop-to-helix packing, enables modeling of the global fold for RNAs up to several hundred nucleotides in length, facilitates ranking of structural models by consistency with through-space restraints, and is poised to enable solution-phase structural interrogation and modeling of complex RNAs. |
| Databáze: | OpenAIRE |
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