Formation of Carboxymethyl-Phosphotriester Adducts in DNA

Area of Science:

• Biochemistry
• Molecular Biology
• Toxicology

Background:

• Humans encounter N-nitroso compounds (NOCs) from endogenous and exogenous sources.
• Metabolic activation of NOCs can produce diazoacetate, linked to DNA adducts and gastrointestinal tumors.
• Previous research focused on nucleobase adducts, neglecting the DNA phosphate backbone.

Purpose of the Study:

• To investigate carboxymethylation on the DNA phosphate backbone.
• To synthesize and characterize carboxymethyl phosphotriester (CM-PTE) adducts.
• To assess the formation of CM-PTE adducts in DNA treated with diazoacetate.

Main Methods:

• Synthesis of a thymidine phosphoramidite building block for CM-PTE.
• Preparation of oligodeoxyribonucleotides (ODNs) with site-specific CM-PTE insertion.
• Liquid-chromatography-tandem mass spectrometry (LC-MS/MS) for adduct identification.

Main Results:

• Successfully synthesized CM-PTE phosphoramidite building blocks.
• Created ODNs containing site-specifically inserted CM-PTE.
• Identified 16 distinct CM-PTE adducts in diazoacetate-treated calf thymus DNA.

Conclusions:

• Carboxymethylation occurs on the DNA phosphate backbone, forming CM-PTE adducts.
• Established methods for synthesizing and detecting CM-PTE lesions.
• Provides a basis for studying in vivo CM-PTE formation and its biological impact.