Unveiling the intricacy of gapmer oligonucleotides through advanced tandem mass spectrometry approaches and scan accumulation for 2DMS,Analyst

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Unveiling the intricacy of gapmer oligonucleotides through advanced tandem mass spectrometry approaches and scan accumulation for 2DMS
Analyst ( IF 3.6 ) Pub Date : 2024-07-18 , DOI: 10.1039/d4an00484a
Mohammed Rahman _{1,

2} , Bryan P Marzullo ₁ , Pui Yiu Lam ₁ , Mark P Barrow ₁ , Stephen W Holman ₃ , Andrew D Ray ₄ , Peter B O'Connor ₁

Affiliation

Antisense oligonucleotides (ASOs) are crucial for biological applications as they bind to complementary RNA sequences, modulating protein expression. ASOs undergo synthetic modifications like phosphorothioate (PS) backbone and locked nucleic acid (LNA) to enhance stability and specificity. Tandem mass spectrometry (MS) techniques were employed to study gapmer ASOs, which feature a DNA chain within RNA segments at both termini, revealing enhanced cleavages with ultraviolet photodissociation (UVPD) and complementary fragment ions from collision-induced dissociation (CID) and electron detachment dissociation (EDD). 2DMS, a data-independent analysis technique, allowed for comprehensive coverage and identification of shared fragments across multiple precursor ions. EDD fragmentation efficiency correlated with precursor ion charge states, with higher charges facilitating dissociation due to intramolecular repulsions. An electron energy of 22.8 eV enabled electron capture and radical-based cleavage. Accumulating multiple scans and generating average spectra improved signal intensity, aided by denoising algorithms. Data analysis utilised a custom Python script capable of handling modifications and generating unique mass lists.

中文翻译：

通过先进的串联质谱方法和 2DMS 扫描累积揭示间隙聚体寡核苷酸的复杂性

反义寡核苷酸 (ASO) 对于生物应用至关重要，因为它们与互补 RNA 序列结合，调节蛋白质表达。 ASO 经过硫代磷酸酯 (PS) 主链和锁核酸 (LNA) 等合成修饰，以增强稳定性和特异性。采用串联质谱 (MS) 技术来研究 gapmer ASO，其特征是两端 RNA 片段内都有 DNA 链，揭示了紫外光解离 (UVPD) 以及碰撞诱导解离 (CID) 和电子脱离产生的互补片段离子的增强裂解解离（EDD）。 2DMS 是一种独立于数据的分析技术，可以全面覆盖和识别多个前体离子的共享碎片。 EDD 碎片效率与前体离子电荷状态相关，较高的电荷由于分子内排斥而促进解离。 22.8 eV 的电子能量能够实现电子捕获和基于自由基的裂解。在去噪算法的帮助下，累积多次扫描并生成平均光谱可提高信号强度。数据分析使用能够处理修改并生成独特质量列表的自定义 Python 脚本。

更新日期：2024-07-18

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