Terminal Alkyne-Modified DNA Aptamers with Enhanced Protein Binding Affinities

Area of Science:

• Biochemistry
• Molecular Biology
• Drug Discovery

Background:

• Aptamers are nucleic acid-based receptors offering rapid discovery and manufacturing.
• Protein receptors (e.g., antibodies) possess diverse functional groups, which aptamers lack.
• Abiotic functional groups can enhance DNA aptamer binding affinity, with aromatic groups like naphthalene showing promise.

Observation:

• Terminal alkynes, known for π-electron-rich interactions in small molecule drugs, were unexplored for aptamer binding enhancement.
• A library of 256 terminal-alkyne variants of the HD22 DNA aptamer (thrombin binder) was synthesized.
• A one-step selection identified a high-affinity aptamer with two alkyne modifications.

Findings:

• The alkyne-modified aptamer exhibited 3.2-fold tighter binding to thrombin compared to the unmodified sequence.
• This enhanced affinity was primarily due to a 5.2-fold slower dissociation rate from thrombin.
• Molecular dynamics simulations revealed π-electron interactions between alkynes and thrombin's asparagine residue, stabilizing the complex.

Implications:

• This study is the first to demonstrate terminal alkynes enhancing aptamer binding properties.
• Terminal alkynes serve as atom-economical, π-electron-rich functional groups for improving aptamer affinity with minimal steric hindrance.
• This finding opens new avenues for designing high-affinity aptamers for therapeutic and diagnostic applications.