Controlling DNA Fragmentation in MALDI-MS by Chemical Modification.

Area of Science:

• Analytical Chemistry
• Biochemistry
• Molecular Biology

Background:

• Fragmentation limits DNA analysis in matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS).
• DNA fragmentation is influenced by the MALDI matrix and nucleic acid sequence.
• Fragmentation involves nucleobase protonation, N-glycosidic bond cleavage, and phosphodiester backbone cleavage.

Purpose of the Study:

• To investigate how structural changes in cytidine analogs affect N-glycosidic bond stability.
• To identify modifications that reduce DNA fragmentation during MALDI-MS.
• To enhance the accessible mass range, sensitivity, and mass resolution of DNA analysis.

Main Methods:

• Synthesized asymmetric oligonucleotides with cytidine analogs (5-methyl-2'-deoxycytidine, 5-bromo-2'-deoxycytidine, aracytidine, 2'-fluorodeoxycytidine).
• Systematically analyzed the impact of nucleobase and deoxyribose sugar modifications on N-glycosidic bond stability.
• Evaluated the effect of 2'-hydroxyl and 2'-fluoro groups on DNA fragmentation.

Main Results:

• Modifications on the deoxyribose sugar ring (2'-hydroxyl, 2'-fluoro) significantly stabilize the N-glycosidic bond.
• 2'-Hydroxyl and 2'-fluoro groups partially or completely inhibit fragmentation at modified nucleosides.
• Oligonucleotides with these modifications showed extended mass range, increased sensitivity, and improved mass resolution.
• Expanded the range of suitable matrices for nucleic acid analysis.

Conclusions:

• Strategic chemical modifications of DNA, particularly at the 2' position of the deoxyribose sugar, effectively suppress fragmentation.
• These stabilized DNA molecules enable superior MALDI-MS performance, including enhanced mass range, sensitivity, and resolution.
• The findings facilitate more robust and comprehensive DNA analysis using MALDI-MS.