APE1 Activity is Controlled by Non-G-Quadruplex Conformations in Single- and Double-Stranded G-Quadruplex Constructs

Area of Science:

• Biochemistry
• Molecular Biology
• DNA Repair

Background:

• Apurinic/apyrimidinic endonuclease-1 (APE1) is a key DNA repair enzyme.
• APE1 typically cleaves abasic (AP) sites in duplex DNA.
• Previous studies indicated reduced APE1 activity on single-stranded G-quadruplexes (ssG4).

Purpose of the Study:

• To investigate the in vitro activity of APE1 on noncanonical DNA structures, specifically G-quadruplexes.
• To determine how G-quadruplex folding affects APE1 endonuclease activity.
• To clarify the cleavage efficiency of APE1 on non-G4 conformations within G4-like structures.

Main Methods:

• Circular dichroism (CD) spectroscopy to analyze DNA structure.
• In vitro activity assays to measure APE1 cleavage rates.
• Studies on various scaffolds including AP-containing ssG4 and duplex-embedded G4 (DGD).

Main Results:

• APE1 efficiently cleaves AP sites in noncanonical, non-G4 conformations of G4-like structures.
• Cleavage yields were comparable to those on duplex DNA substrates.
• APE1 activity significantly decreased with increasing G-quadruplex folding in both ssG4 and DGD systems.
• A positional dependency of cleavage yield was observed in non-G4 DGD scaffolds.

Conclusions:

• APE1 demonstrates efficient cleavage of noncanonical DNA conformations.
• The secondary structure of DNA, particularly G-quadruplex folding, critically controls APE1 endonuclease activity.
• This highlights the enzyme's ability to act on structurally diverse DNA substrates beyond canonical duplexes.