Wan Jiahao, Nytka Marianna, Qian Haocheng, Lemr Karel, Tureček František
Department of Chemistry, University of Washington, Bagley Hall, Box 351700, Seattle, Washington 98195-1700, United States.
Department of Analytical Chemistry, Faculty of Science, Palacký University, 17. listopadu 12, 779 00 Olomouc, Czech Republic.
J Am Soc Mass Spectrom. 2023 Oct 4;34(10):2323-2340. doi: 10.1021/jasms.3c00228. Epub 2023 Sep 11.
d(GCGAAGC) is the smallest oligonucleotide with a well-defined hairpin structure in solution. We report a study of multiply protonated d(GCGAAGC) and its sequence-scrambled isomers, d(CGAAGCG), d(GCGAACG), and d(CGGAAGC), that were produced by electrospray ionization with the goal of investigating their gas-phase structures and dissociations. Cyclic ion mobility measurements revealed that dications of d(GCGAAGC) as well as the scrambled-sequence ions were mixtures of protomers and/or conformers that had collision cross sections (CCS) within a 439-481 Å range. Multiple ion conformers were obtained by electrospray under native conditions as well as from aqueous methanol. Arrival time distribution profiles were characteristic of individual isomeric heptanucleotides. Extensive Born-Oppenheimer molecular dynamics (BOMD) and density functional theory (DFT) calculations of d(GCGAAGC) isomers indicated that hairpin structures were high-energy isomers of more compact distorted conformers. Protonation caused a break up of the C2···G6 pair that was associated with the formation of strong hydrogen bonds in zwitterionic phosphate anion-nucleobase cation motifs that predominated in low energy ions. Multiple components were also obtained for d(GCGAAGC) trications under native and denaturing electrospray conditions. The calculated trication structures showed disruption of the G···C pairs in low energy zwitterions. A hairpin trication was calculated to be a high energy isomer. d(GCGAAGC) tetracations were produced and separated by c-IMS as two major isomers. All low energy d(GCGAAGC) ions obtained by DFT geometry optimizations were zwitterions in which all five purine bases were protonated, and the ion charge was balanced by a phosphate anion. Tetracations of the scrambled sequences were each formed as one dominant isomer. The CCS calculated with the MobCal-MPI method were found to closely match experimental values. Collision-induced dissociation (CID) spectra of multiply charged heptanucleotides showed nucleobase loss and backbone cleavages occurring chiefly at the terminal nucleosides. Electron-transfer-CID tandem mass spectra were used to investigate dissociations of different charge and spin states of charge-reduced heptanucleotide cation radicals.
d(GCGAAGC)是溶液中具有明确发夹结构的最小寡核苷酸。我们报告了一项对多重质子化的d(GCGAAGC)及其序列打乱的异构体d(CGAAGCG)、d(GCGAACG)和d(CGGAAGC)的研究，这些异构体是通过电喷雾电离产生的，目的是研究它们的气相结构和解离情况。循环离子迁移率测量表明，d(GCGAAGC)的双电荷离子以及序列打乱的离子是原聚体和/或构象异构体的混合物，其碰撞截面（CCS）在439 - 481 Å范围内。在天然条件下以及从甲醇水溶液中通过电喷雾获得了多个离子构象异构体。到达时间分布曲线是各个异构七核苷酸的特征。对d(GCGAAGC)异构体进行的广泛的玻恩 - 奥本海默分子动力学（BOMD）和密度泛函理论（DFT）计算表明，发夹结构是更紧凑扭曲构象异构体的高能异构体。质子化导致C2···G6碱基对断裂，这与两性离子磷酸根阴离子 - 核碱基阳离子基序中强氢键的形成有关，这种基序在低能离子中占主导。在天然和变性电喷雾条件下，d(GCGAAGC)的三电荷离子也获得了多个组分。计算得到的三电荷离子结构显示低能两性离子中G···C碱基对的破坏。计算得出发夹三电荷离子是一种高能异构体。d(GCGAAGC)的四电荷离子通过c - IMS产生并分离为两种主要异构体。通过DFT几何优化获得的所有低能d(GCGAAGC)离子都是两性离子，其中所有五个嘌呤碱基都被质子化，离子电荷由磷酸根阴离子平衡。序列打乱的异构体的四电荷离子各自形成为一种主要异构体。发现用MobCal - MPI方法计算的CCS与实验值密切匹配。多重带电七核苷酸的碰撞诱导解离（CID）光谱显示，碱基损失和主链断裂主要发生在末端核苷处。电子转移 - CID串联质谱用于研究电荷减少的七核苷酸阳离子自由基不同电荷和自旋态的解离情况。
