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Paranemic Cohesion of DNA under Isothermal Conditions.

Lauren A Anderson1, Akul Patel1, Bharath Raj Madhanagopal1

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This study demonstrates the isothermal assembly of paranemic crossover (PX) DNA motifs using various divalent metal ions. This method simplifies DNA nanotechnology and allows for the attachment of biomolecules like aptamers at constant temperatures.

Keywords:
DNA nanostructuresDNA nanotechnologyPX DNAcounterionsisothermal assemblyparanemic crossover DNA

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Area of Science:

  • DNA nanotechnology
  • Biomolecular engineering
  • Materials science

Background:

  • Isothermal assembly simplifies DNA nanostructure construction, avoiding thermal cyclers and enabling biomolecule attachment at ambient temperatures.
  • The paranemic crossover (PX) DNA motif is utilized in DNA nanostructure construction and may play a role in homology recognition.
  • Previous methods often required thermal cycling, limiting applications and biomolecule compatibility.

Purpose of the Study:

  • To demonstrate the successful isothermal assembly of the PX DNA motif in the presence of different divalent metal ions (Mg2+, Ca2+, Sr2+).
  • To investigate the thermodynamic properties of PX DNA interhelix hybridization.
  • To showcase the utility of isothermal assembly for site-specific attachment of functional biomolecules, such as aptamers.

Main Methods:

  • Isothermal assembly of PX DNA motifs at constant temperatures (20 and 37 °C) in solutions containing Mg2+, Ca2+, and Sr2+.
  • Isothermal titration calorimetry (ITC) to determine thermodynamic parameters of hybridization.
  • Design and assembly of PX DNA containing thrombin-specific aptamers.

Main Results:

  • Successful isothermal assembly of PX DNA motifs was achieved in Mg2+, Ca2+, and Sr2+ at both tested temperatures.
  • Isothermal titration calorimetry revealed that interhelix hybridization of half-PX molecules is thermodynamically favored at higher temperatures (ΔCp = -1.9 kcal/mol·K).
  • PX DNA nanostructures functionalized with thrombin-specific aptamers were assembled isothermally, demonstrating site-specific binding of thrombin molecules.

Conclusions:

  • Isothermal assembly is a versatile method for constructing complex DNA motifs like PX DNA in various ionic conditions.
  • The study extends the application of isothermal assembly to include different counterions and complex DNA structures.
  • This work highlights the potential for isothermal attachment of guest biomolecules to DNA nanostructures for diverse applications in nanotechnology and synthetic biology.