The trinucleotide repeat sequence d(GTC)15 adopts a hairpin conformation

Area of Science:

• Molecular Biology
• Biophysics
• Oligonucleotide Chemistry

Background:

• Single-stranded oligonucleotides can adopt complex secondary structures.
• Hairpin formation is a common structural motif in nucleic acids.
• Understanding oligonucleotide structure is crucial for applications in molecular biology and therapeutics.

Purpose of the Study:

• To investigate and compare the hairpin structures formed by single-stranded (GTC)15 and (CTG)15 oligonucleotides.
• To elucidate the structural differences and their implications for stability and nuclease recognition.

Main Methods:

• Electrophoretic mobility studies to assess structure and melting temperatures.
• Chemical probing using KMnO4 oxidation to identify accessible bases.
• Enzymatic digestion with single-strand-specific P1 nuclease.
• Molecular dynamics simulations to model hairpin conformation.

Main Results:

• Both ss(GTC)15 and ss(CTG)15 form hairpin structures with base-paired or stacked thymines in the stem.
• Melting temperatures were 38°C for ss(GTC)15 and 48°C for ss(CTG)15.
• P1 nuclease cleaved distinct phosphodiester bonds in the loops, indicating differential accessibility.
• Molecular dynamics revealed loop bending in ss(GTC)15 and base separation in ss(CTG)15 due to guanine stacking.

Conclusions:

• ss(GTC)15 and ss(CTG)15 form similar yet distinguishable hairpin structures.
• Structural variations arise from loop conformation and base stacking interactions.
• These structural nuances influence stability and susceptibility to enzymatic cleavage.