Nitric oxide-induced interstrand cross-links in DNA

Area of Science:

• Biochemistry
• Molecular Biology
• Genetics

Background:

• Nitrous acid is known to induce DNA interstrand cross-links.
• Nitric oxide (NO) is a biological molecule that can generate nitrosating agents.
• The formation of DNA cross-links via nitric oxide is not well understood.

Purpose of the Study:

• To investigate the potential for DNA interstrand cross-linking mediated by nitric oxide.
• To quantify the formation of interstrand cross-links induced by nitric oxide in a specific DNA oligomer.
• To compare the yield of cross-links to the yield of a known deamination product.

Main Methods:

• Utilized a high-performance liquid chromatography (HPLC) laser-induced fluorescence (LIF) system for cross-link detection.
• Employed gas chromatography-mass spectrometry (GC/MS) to quantify the deamination product, xanthine.
• Treated a 5'-fluorescein-labeled oligomer (ATATCGATCGATAT) containing CpG sites with nitric oxide under controlled conditions.

Main Results:

• Nitric oxide treatment of the oligomer resulted in the formation of interstrand cross-links.
• The concentration of interstrand cross-links was approximately 6% of the xanthine concentration (13 +/- 2.5 nM cross-links vs. 211 +/- 39 nM xanthine).
• A quasi-linear response was observed between nitric oxide concentration and cross-link formation, with lower doses yielding detectable cross-links.

Conclusions:

• Nitric oxide can indeed induce DNA interstrand cross-links, likely through a nitrosating intermediate.
• The study provides a method for quantifying these cross-links using HPLC-LIF.
• Findings suggest the possibility of predicting genome-wide cross-links based on CpG content and nitrosative deamination yields.