Defining the substrate scope of DNAzyme catalysis for reductive amination with aliphatic amines

Area of Science:

• Biochemistry
• Molecular Biology
• Catalysis

Background:

• Amines undergo alkylation via reactions like reductive amination.
• DNAzymes, catalytic DNA sequences, offer potential for novel chemical transformations.
• Understanding DNAzyme substrate scope is crucial for expanding their catalytic applications.

Purpose of the Study:

• To identify DNAzymes capable of catalyzing reductive amination with aliphatic amines.
• To investigate the substrate scope limitations of DNAzyme-catalyzed N-alkylation.
• To explore DNAzyme activity with DNA-anchored peptide substrates containing lysine.

Main Methods:

• In vitro selection using N40 and N20 random DNA pools.
• Reductive amination assays with DNA-anchored substrates and benzaldehyde.
• Kinetic analysis under varying pH and metal ion cofactor conditions.
• Analogous selection experiments with complex peptide substrates.

Main Results:

• Identified DNAzymes catalyzing reductive amination of DNA-C3-NH2 with benzaldehyde, showing up to 130-fold rate enhancement.
• DNAzymes were not found for reductive amination with longer tethered amines or lysine-containing peptide substrates.
• No DNAzymes were identified even when other amino acids replaced neighboring alanines in peptide substrates.

Conclusions:

• DNAzyme catalysis for N-alkylation via reductive amination has a practical substrate scope limit.
• The complexity of binding and spatial organization of substrates challenges DNAzyme development for more complex molecules.
• DNAzyme identification is highly dependent on the specific reaction and substrate composition.