An Efficient Catalytic DNA that Cleaves L-RNA.

Area of Science:

• Biochemistry
• Molecular Biology
• Biotechnology

Background:

• DNAzymes cleave natural RNA (D-RNA) substrates and are used in aptazyme biosensors.
• Existing biosensors fail in complex biological samples due to RNase degradation of D-RNA substrates.

Purpose of the Study:

• To develop DNAzymes capable of cleaving L-RNA, which is resistant to RNases.
• To engineer robust aptazyme biosensors for detecting analytes in complex biological matrices.

Main Methods:

• In vitro selection was employed to isolate L-RNA-cleaving DNAzymes from a DNA pool.
• The catalytic activity of the isolated DNAzymes was characterized.
• An aptazyme sensor was constructed using a novel L-RNA-cleaving DNAzyme and an ATP-binding aptamer.

Main Results:

• Three L-RNA-cleaving DNAzymes were successfully isolated.
• The most active DNAzyme demonstrated a catalytic rate constant of approximately 3 min⁻¹.
• This DNAzyme featured a unique kissing loop structural motif.
• The developed aptazyme biosensor successfully detected ATP in RNase-containing biological samples.

Conclusions:

• Novel L-RNA-cleaving DNAzymes were discovered, offering resistance to RNase degradation.
• These DNAzymes enable the development of biosensors compatible with complex biological samples.
• The findings pave the way for engineering advanced RNA-cleaving DNAzymes for biosensing applications.