Recent interest in biological and synthetic DNA nanostructures has highlighted the need for methods to comprehensively characterize intermediates and end products of multimeric DNA assembly. Here we use native mass spectrometry in combination with ion mobility to determine the mass, charge state and collision cross section of noncovalent DNA assemblies, and thereby elucidate their structural composition, oligomeric state, overall size and shape. We showcase the approach with a prototypical six-subunit DNA nanostructure to reveal how its assembly is governed by the ionic strength of the buffer, as well as how the mass and mobility of heterogeneous species can be well resolved by careful tuning of instrumental parameters. We find that the assembly of the hexameric, barrel-shaped complex is guided by positive cooperativity, while previously undetected higher-order 12- and 18-mer assemblies are assigned to defined larger-diameter geometric structures. Guided by our insight, ion mobility-mass spectrometry is poised to make significant contributions to understanding the formation and structural diversity of natural and synthetic oligonucleotide assemblies relevant in science and technology.

最近对生物和合成 DNA 纳米结构的兴趣凸显了对全面表征多聚体 DNA 组装的中间体和最终产物的方法的需求。在这里，我们使用天然质谱法结合离子淌度来确定非共价 DNA 组装体的质量、电荷状态和碰撞截面，从而阐明它们的结构组成、寡聚状态、总体大小和形状。我们展示了具有原型六亚基 DNA 纳米结构的方法，以揭示其组装如何受缓冲液离子强度的控制，以及如何通过仔细调整仪器参数来很好地解决异质物种的质量和流动性。我们发现六聚体、桶形复合体的组装是由正协同性引导的，而以前未检测到的高阶 12 和 18 聚体组装被分配给定义的较大直径几何结构。在我们的洞察力的指导下，离子淌度-质谱法有望为理解与科学技术相关的天然和合成寡核苷酸组装的形成和结构多样性做出重大贡献。