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bioRxiv · 10.1101/2025.07.29.667413

Native Ion Mobility-MS Reveals the Bead Ejection Mechanism in Multi-Domain Nucleic Acids

Ghosh, D.·Rosu, F.·Gabelica, V.
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Abstract

Understanding how biomolecules acquire their charge and retain their solution conformation during electrospray ionization (ESI) is crucial for native mass spectrometry (native MS) interpretation. Here, we examine the charging and gas phase conformation of nucleic acid constructs comprising folded G-quadruplex "beads" linked by unstructured polythymine regions. Under physiological ionic strength, these oligonucleotides exhibit a multimodal charge-state and collision cross section distribution, revealing multiple conformational ensembles, in contrast to the unimodal profiles typically observed for shorter oligonucleotides. Native MS observations for intermediate charge states are compatible with ion production via the recently proposed bead ejection scenario, in addition to the charge residue scenario for low charge states and chain ejection for the highest charge states or for sequences with thymine overhangs on both ends. The preservation of the local structures in ions charged above the Rayleigh limit helps to infer the presence of folded subunits. The position of the G-quadruplex subunit and ionic strength governs the charging and retention of G-quadruplex folded regions. Our findings broaden the existing conceptual framework underpinning nucleic acid ionization. TOC graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=167 SRC="FIGDIR/small/667413v2_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@42d7c3org.highwire.dtl.DTLVardef@63445dorg.highwire.dtl.DTLVardef@11d7056org.highwire.dtl.DTLVardef@1ba48bb_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Ghosh, D., Rosu, F., Gabelica, V.. 2025-08-01. Native Ion Mobility-MS Reveals the Bead Ejection Mechanism in Multi-Domain Nucleic Acids. https://doi.org/10.1101/2025.07.29.667413

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