Construction of a G-quadruplex-specific DNA endonuclease

Area of Science:

• Biochemistry
• Molecular Biology
• Genomics

Background:

• G-quadruplexes (G4s) are non-canonical DNA secondary structures implicated in various biological processes.
• Detecting and mapping G4s in complex genomic DNA remains a challenge.

Purpose of the Study:

• To develop a novel endonuclease with specificity for G-quadruplex DNA structures.
• To create a tool for the detection and mapping of G4s within double-stranded DNA (dsDNA).

Main Methods:

• Engineered a fusion protein by combining the G4 recognition domain of the RHAU helicase with the Fok1 nuclease cleavage domain.
• Assessed the binding specificity of the fusion protein to parallel G4 structures.
• Evaluated the cleavage activity of the fusion protein on dsDNA adjacent to G4s.

Main Results:

• Successfully generated a novel G4-specific endonuclease.
• The fusion protein demonstrated specific binding to parallel G4 structures.
• The endonuclease was shown to cleave double-stranded DNA (dsDNA) located next to the bound G4.

Conclusions:

• The novel G4-specific endonuclease is a potent tool for G4 detection.
• This enzyme facilitates the mapping of G4 formation within long dsDNA, including genomic contexts.
• The developed endonuclease advances G4 research and genomic analysis tools.