Boronic Acid Assisted Self-Assembly of Functional RNAs

Area of Science:

• Biochemistry
• Molecular Biology
• Synthetic Biology

Background:

• Oligonucleotides with 5'-boronic acid moieties can form boronate esters with 3'-terminal cis-diols.
• These boronate esters have previously aided in assembling fragmented DNAzymes.
• Functional RNAs like ribozymes and aptamers are sensitive to fragmentation, often losing activity.

Purpose of the Study:

• To investigate the use of boronate esters as internucleoside linkages in functional RNAs.
• To assess the ability of boronate esters to restore function to fragmented RNA molecules.
• To determine if boronate esters can act as surrogates for natural phosphodiester bonds in RNA.

Main Methods:

• Synthesized RNA fragments with 5'-boronic acid moieties.
• Created boronate ester linkages between RNA fragments, replacing phosphodiesters.
• Assessed the functional activity (cleavage, ligation, binding) of assembled RNAs.

Main Results:

• Boronate ester linkages restored cleavage activity in some fragmented hairpin ribozymes, depending on the split site.
• Ligation activity of the hairpin ribozyme was not restored by boronate ester linkages.
• Fragmented Mango aptamers regained functionality when assembled using 5'-boronic acid modified fragments.
• Boronate esters were shown to be viable internucleoside linkages in functional RNA.

Conclusions:

• Boronate esters can serve as effective surrogates for natural phosphodiester linkages in functional RNA molecules.
• The efficacy of boronate ester assembly depends on the specific RNA and the site of fragmentation.
• This work opens new avenues for RNA assembly and engineering using non-natural linkages.