Chemistry Department and GIGA-R, Mass Spectrometry Laboratory, University of Liege, Liege, Belgium.
J Am Soc Mass Spectrom. 2013 Nov;24(11):1777-86. doi: 10.1007/s13361-013-0721-y. Epub 2013 Sep 6.
Electrospray ionization (ESI) soft desolvation is widely used to investigate fragile species such as nucleic acids. Tandem mass spectrometry (MS/MS) gives access to the gas phase energetics of the intermolecular interactions in the absence of solvent, by following the dissociation of mass-selected ions. Ion mobility mass spectrometry (IMS) provides indications on the tridimensional oligonucleotide structure by attributing a collision cross section (CCS) to the studied ion. Electrosprayed duplexes longer than eight bases pairs retain their helical structure in a solvent-free environment. However, the question of conformational changes under activation in MS/MS studies remains open. The objective of this study is to probe binding energetics and characterize the unfolding steps occurring prior to oligonucleotide duplex dissociation. Comparing the evolution of CCS with collision energy and breakdown curves, we characterize dissociation pathways involved in CID-activated DNA duplex separation into single strands, and we demonstrate here the existence of stable dissociation intermediates. At fixed duplex length, dissociation pathways were found to depend on the percentage of GC base pairs and on their position in the duplex. Our results show that pure GC sequences undergo a gradual compaction until reaching the dissociation intermediate: A-helix. Mixed AT-GC sequences were found to present at least two conformers: a classic B-helix and an extended structure where the GC tract is a B-helix and the AT tract(s) fray. The dissociation in single strands takes place from both conformers when the AT base pairs are enclosed between two GC tracts or only from the extended conformer when the AT tract is situated at the end(s) of the sequence.
电喷雾电离（ESI）软解吸广泛用于研究核酸等脆弱物种。串联质谱（MS/MS）通过跟踪质量选择离子的解离，在没有溶剂的情况下获得分子间相互作用的气相能量学信息。离子淌度质谱（IMS）通过将碰撞截面（CCS）分配给研究离子，提供有关三维寡核苷酸结构的信息。在无溶剂环境中，超过八个碱基对的电喷雾双链体保留其螺旋结构。然而，在 MS/MS 研究中关于构象变化的问题仍然存在。本研究的目的是探测结合能，并表征在寡核苷酸双链体解离之前发生的展开步骤。通过比较 CCS 与碰撞能和分解曲线的演变，我们表征了在 CID 激活的 DNA 双链体分离成单链过程中涉及的解离途径，并在此证明了稳定解离中间体的存在。在固定的双链体长度下，发现解离途径取决于 GC 碱基对的百分比及其在双链体中的位置。我们的结果表明，纯 GC 序列经历逐渐的紧缩，直到达到解离中间体：A 螺旋。混合 AT-GC 序列被发现至少存在两种构象：经典 B 螺旋和扩展结构，其中 GC 链段为 B 螺旋，AT 链段（s）散开。当 AT 碱基对被两个 GC 链段包围时，双链体从两种构象解离成单链，而当 AT 链段位于序列的末端时，只从扩展构象解离成单链。
