How long is too long? Effects of loop size on G-quadruplex stability

Area of Science:

• Biochemistry
• Molecular Biology
• Oligonucleotide Chemistry

Background:

• Guanine-rich sequences can form stable G-quadruplex structures.
• The stability of G-quadruplexes is affected by loop length and cation concentration.
• Current sequence design often uses a maximal loop size consensus.

Purpose of the Study:

• To investigate the impact of variable loop lengths on G-quadruplex stability.
• To compare G-quadruplex stability in potassium (K+) versus sodium (Na+) solutions.
• To expand the repertoire of sequences capable of forming stable G-quadruplexes.

Main Methods:

• Synthesized and analyzed 80 different G-rich oligonucleotide sequences.
• Measured melting temperatures (Tm) to assess G-quadruplex stability.
• Varied loop lengths from 1 to 30 bases and tested in K+ and Na+ solutions.

Main Results:

• All tested sequences formed stable G-quadruplexes with Tm above physiological temperature.
• G-quadruplex stability was consistently lower in Na+ than K+, with significant variations.
• A strong inverse correlation between total loop length and Tm was observed in K+.
• Optimal loop length in Na+ was longer than anticipated, up to 5 nucleotides.

Conclusions:

• Loop length significantly impacts G-quadruplex stability, with an inverse relationship in K+.
• Ionic conditions (K+ vs. Na+) markedly affect G-quadruplex stability and optimal loop length.
• The findings support expanding the sequence space for G-quadruplex formation and suggest revising the 7-base loop size consensus.