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Probing The Structure And Dynamics Of Nucleosomes Using Atomic Force Microscopy Imaging
Published on: January 31, 2019
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Visualizing the functional 3D shape and topography of long noncoding RNAs by single-particle atomic force microscopy
Tina Uroda1,2, Isabel Chillón1, Paolo Annibale3
1European Molecular Biology Laboratory (EMBL) Grenoble, Grenoble, France.
Nature Protocols
|May 27, 2020
Summary
Researchers developed a new protocol to visualize the 3D structures of long noncoding RNAs (lncRNAs) using atomic force microscopy. This method enables high-resolution structural characterization of these complex molecules, crucial for understanding their cellular roles and disease connections.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- Long noncoding RNAs (lncRNAs) are vital regulators of cellular processes and disease.
- The complex 3D structures of lncRNAs are critical for their function but challenging to determine.
- High-resolution structural characterization of lncRNAs is essential for understanding their roles.
Purpose of the Study:
- To present a novel protocol for the biochemical and biophysical characterization of lncRNAs.
- To enable high-resolution 3D structural visualization of lncRNAs.
- To facilitate the study of lncRNA structure-function relationships and disease mechanisms.
Main Methods:
- Integration of non-denaturing lncRNA purification with in-solution hydrodynamic analysis.
- Single-particle atomic force microscopy (AFM) imaging for 3D topology visualization.
- Protocol adaptable for biologically active conformations and mutant analysis.
Main Results:
- Production of highly homogeneous lncRNA preparations.
- Visualization of lncRNA 3D topology at approximately 15-Å resolution.
- Protocol suitable for analyzing structural defects in functionally inactive mutants.
Conclusions:
- The developed protocol provides a powerful approach for high-resolution lncRNA structural characterization.
- This method allows imaging of lncRNAs in biologically relevant conformations.
- The protocol aids in understanding lncRNA function, regulation, and disease association.
