DNA-Catalyzed Introduction of Azide at Tyrosine for Peptide Modification

Area of Science:

• Biochemistry
• Molecular Biology
• Synthetic Biology

Background:

• DNA enzymes (deoxyribozymes) are catalytic DNA molecules with diverse applications.
• Chemical modification of peptides is crucial for developing new therapeutics and research tools.
• Targeting specific amino acid residues like tyrosine for modification remains a challenge.

Purpose of the Study:

• To develop novel deoxyribozymes capable of introducing azide functional groups onto tyrosine residues in peptide substrates.
• To explore both general and sequence-selective deoxyribozymes for this modification.
• To demonstrate the utility of these modified peptides by attaching polyethylene glycol (PEG) and fluorescent labels.

Main Methods:

• In vitro selection was employed to identify and evolve deoxyribozymes with the desired catalytic activity.
• The deoxyribozymes were designed to transfer a specific azido-containing nucleotide group (2'-Az-dAMP) from its triphosphate precursor (2'-Az-dATP).
• Peptide substrates were either tethered to a DNA anchor or used as free molecules during the enzymatic reaction.

Main Results:

• Novel deoxyribozymes were successfully identified that catalyze the azidation of tyrosine residues in peptides.
• The developed deoxyribozymes demonstrated both general and sequence-selective activity, allowing for versatile applications.
• The azido-modified peptides were further functionalized with PEG moieties and fluorescent labels using the engineered deoxyribozymes.

Conclusions:

• Deoxyribozyme-mediated azidation of tyrosine residues in peptides is a feasible and powerful strategy.
• This methodology provides a novel tool for site-specific peptide modification.
• The ability to attach various moieties like PEG and fluorescent labels opens avenues for bioconjugation and chemical biology research.