Trapping Transient RNA Complexes by Chemically Reversible Acylation.

Area of Science:

• Molecular Biology
• Biochemistry
• RNA Biology

Background:

• RNA-RNA interactions are crucial in biological processes but challenging to study due to their transient nature.
• Existing RNA crosslinking methods are often irreversible, chemically altering RNA and impeding downstream molecular analyses.

Purpose of the Study:

• To develop a novel, reversible crosslinking strategy for studying RNA-RNA interactions.
• To enable efficient capture and subsequent analysis of transient RNA complexes.

Main Methods:

• Introduction of a novel soluble crosslinker, BINARI, which reacts with RNA via acylation.
• Demonstration of BINARI's ability to efficiently crosslink noncovalent RNA complexes with minimal sequence bias.
• Establishment of reversible crosslink cleavage using phosphine reduction of azide trigger groups.

Main Results:

• BINARI efficiently crosslinks RNA complexes with low sequence specificity.
• The BINARI crosslink is reversible, allowing for the liberation of individual RNA molecules.
• Demonstrated utility in reversible protection against nuclease degradation and trapping of transient RNA complexes, such as E. coli DsrA-rpoS bulge-loop interactions.

Conclusions:

• BINARI provides a valuable tool for studying transient RNA-RNA interactions.
• The reversibility of BINARI crosslinking facilitates further molecular analysis of captured RNA complexes.
• This approach has significant potential for investigating complex RNA regulatory networks.