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Direct Mapping of Higher-Order RNA Interactions by SHAPE-JuMP
Thomas W Christy1,2, Catherine A Giannetti1, Gillian Houlihan3
1Department of Chemistry, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-3290, United States.
Biochemistry
|June 14, 2021
Summary
We developed SHAPE-JuMP to map RNA tertiary structures in solution. This method uses chemical cross-linking and a novel reverse transcriptase to detect nucleotide proximity, aiding complex RNA structure analysis.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- RNA higher-order structure dictates function.
- Determining large RNA structures in solution remains challenging.
Purpose of the Study:
- To introduce SHAPE-JuMP (selective 2 ahydroxyl acylation analyzed by primer extension and juxtaposed merged pairs) for interrogating through-space RNA tertiary interactions.
- To provide a method for analyzing complex RNA structures in solution.
Main Methods:
- Utilizes a bifunctional small molecule to cross-link proximal nucleotides in RNA.
- Employs an engineered reverse transcriptase to "jump" across cross-links, encoding them into cDNA.
- Detects cross-linked sites via sequencing, identifying deletions in the resulting cDNA.
Main Results:
- SHAPE-JuMP measures RNA tertiary structure proximity at nanometer resolution.
- Effective for interactions in multihelix junctions and loop-to-helix packing.
- Enables global fold modeling for RNAs up to several hundred nucleotides.
Conclusions:
- SHAPE-JuMP facilitates the ranking of structural models based on through-space restraints.
- Poised to enable solution-phase structural interrogation and modeling of complex RNAs.
- Advances the understanding of RNA structure-function relationships.
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