Ion mobility spectrometry reveals duplex DNA dissociation intermediates

Area of Science:

• Biophysical Chemistry
• Mass Spectrometry
• Structural Biology

Background:

• Electrospray ionization (ESI) and tandem mass spectrometry (MS/MS) are crucial for studying fragile biomolecules like nucleic acids in the gas phase.
• Ion mobility mass spectrometry (IMS) provides insights into oligonucleotide 3D structures via collision cross-section (CCS) measurements.
• While duplexes longer than eight base pairs maintain helical structure post-ESI, conformational changes during MS/MS activation remain unclear.

Purpose of the Study:

• To investigate the binding energetics of DNA duplexes.
• To characterize the unfolding steps preceding oligonucleotide duplex dissociation.
• To elucidate conformational changes and dissociation pathways under activation in MS/MS.

Main Methods:

• Utilizing ion mobility mass spectrometry (IMS) to measure collision cross sections (CCS).
• Employing tandem mass spectrometry (MS/MS) with collision-induced dissociation (CID) to activate and dissociate DNA duplex ions.
• Analyzing the correlation between CCS, collision energy, and breakdown curves to map dissociation pathways.

Main Results:

• Stable dissociation intermediates were identified during CID-activated DNA duplex separation into single strands.
• Dissociation pathways are influenced by the percentage and position of GC base pairs within the duplex.
• Pure GC sequences compact gradually to an A-helix intermediate, while mixed AT-GC sequences exhibit B-helix and extended fraying conformers.

Conclusions:

• The study demonstrates stable intermediates in DNA duplex dissociation, offering new insights into gas-phase structural dynamics.
• Conformational heterogeneity in mixed AT-GC sequences affects their dissociation behavior, with implications for understanding DNA structural transitions.
• The findings highlight the utility of IMS-MS for probing the energetic and structural aspects of oligonucleotide dissociation.