[论文解读] Hairygami: Analysis of DNA Nanostructures' Conformational Change Driven by Functionalizable Overhangs

Matthew Sample, Hao Liu+3 more|arXiv (Cornell University)|Feb 17, 2023

Advanced biosensing and bioanalysis techniquesBiochemistry, Genetics and Molecular Biology参考文献 49被引用 3

一句话总结

本研究通过oxDNA粗粒化分子动力学模拟并经TEM实验验证，揭示了可功能化的单链DNA悬突在熵效应驱动下诱导2D DNA纳米折纸Tile产生弯曲。该现象源于悬突改变了溶液中的构象分布，挑战了折纸结构保持平面的假设，对设计稳定DNA纳米结构及通过悬突工程调控其形状具有重要意义。

ABSTRACT

DNA origami is a widely used method to construct nanostructures by self-assembling designed DNA strands. These structures are often used as "pegboards" for templated assembly of proteins, gold nanoparticles, aptamers, and other molecules, with applications ranging from therapeutics and diagnostics to plasmonics and photonics. Imaging these structures using AFM or TEM does not capture their full conformation ensemble as they only show their shape flattened on a surface. However, certain conformations of the nanostructure can position guest molecules into distances unaccounted for in their intended design, thus leading to spurious interactions between guest molecules that are designed to be separated. Here, we use molecular dynamics simulations to capture conformational ensemble of 2D DNA origami tiles and show that introducing single-stranded overhangs, which are typically used for functionalization of the origami with guest molecules, induces a curvature of the tile structure in the bulk. We show that the shape deformation is of entropic origin, with implications for design of robust DNA origami breadboards as well as potential approach to modulate structure shape by introducing overhangs. We then verify experimentally that the DNA overhangs introduce curvature into the DNA origami tiles in divalent as well as monovalent salt buffer conditions. We further experimentally verify that DNA origami functionalized with attached proteins also experience such induced curvature. We provide the developed simulation code implementing the enhanced sampling to characterize conformational space of DNA origami as open source software.

研究动机与目标

• 研究可功能化的DNA悬突如何影响溶液中2D DNA纳米折纸Tile的构象动力学。
• 解决基于表面的成像技术（如AFM、TEM）无法捕捉溶液相构象分布的局限性。
• 确定悬突是否在设计意图之外诱导DNA折纸结构发生形变。
• 建立计算与实验框架，用于表征DNA纳米结构在体相溶液中的构象分布。
• 通过理解悬突诱导的形状变化，为设计稳定DNA折纸基板提供设计原则。

提出的方法

• 采用GPU加速的oxDNA粗粒化模型（版本2），模拟具有不同悬突长度的2D DNA纳米折纸Tile。
• 使用伞形采样结合谐振势能，以端到端距离（R_ee）为有序参数计算自由能剖面。
• 应用加权直方图分析法（WHAM）对模拟数据去偏置，并以200个bin重建自由能景观。
• 通过蒙特卡洛自助法对自由能剖面和加权平均值进行误差估计。
• 采用多项式拟合将R_ee与弯曲角度关联，以辅助结构解释。
• 通过凝胶电泳和负染色透射电子显微镜（TEM）实验验证模拟预测。

Figure 1: The addition of overhangs causes a $2$ D DNA origami tile $(a)$ to adopt a curved shape $(b)$ . The origin of the curvature is mainly due to the entropic penalty of the overhang sequences on a flat surface $(c)$ , compared to a curved surface $(d)$ (shown schematically as a side view of an

实验结果

研究问题

• RQ1 在溶液中，可功能化的DNA悬突是否诱导2D DNA纳米折纸Tile发生结构弯曲？
• RQ2 悬突诱导的弯曲其热力学起源是熵效应还是焓效应？
• RQ3 悬突长度与位置如何影响DNA折纸在溶液中的构象分布？
• RQ4 分子动力学模拟能否准确预测带有悬突的DNA折纸在溶液中的形态？
• RQ5 AFM和TEM等基于表面的成像技术在多大程度上无法真实反映DNA折纸在溶液中的构象？

主要发现

• 单链悬突在溶液中可诱导2D DNA纳米折纸Tile产生可测量的弯曲，且弯曲程度随悬突长度增加而增大。
• 弯曲由熵效应驱动，自由能剖面证实弯曲构象具有偏好性。
• 自由能景观显示存在一个稳定的弯曲态，其最小值出现在R_ee ≈ 30 nm处，表明存在偏好弯曲构象。
• 在单张GPU上并行运行40个悬突的模拟，采样效率提升2.6倍，实现了大规模构象分析。
• 实验TEM成像证实了功能化悬突折纸Tile中存在弯曲，验证了模拟预测。
• 本研究证明，悬突可作为设计元件，用于主动调控溶液中DNA纳米结构的形状。

Figure 2: $(a)$ Mean structure of twist-corrected rectangular origami with $169$ overhang extensions comprised of twenty nucleotide bases. The arrows indicate the measured end-to-end distance order parameter ( $R_{\rm ee}$ ) used to model the curvature of the structures. Low $R_{\rm ee}$ values corr