Molecular Insights into the Binding of Linear Polyethylenimines and Single-Stranded DNA Using Raman Spectroscopy: A Quantitative Approach

Establishing how polymeric vectors such as polyethylenimine (PEI) bind and package their nucleic acid cargo is vital toward developing more efficacious and cost-effective gene therapies. To develop a molecular-level picture of DNA binding, we examined how the Raman spectra of PEIs report on their local chemical environment. We find that the intense Raman bands located in the 1400–1500 cm–1 region derive from vibrations with significant CH2 scissoring and NH bending character. The Raman bands that derive from these vibrations show profound intensity changes that depend on both the local dielectric environment and hydrogen bonding interactions with the secondary amine groups on the polymer. We use these bands as spectroscopic markers to assess the binding between low molecular weight PEIs and single-stranded DNA (ssDNA). Analysis of the Raman spectra suggest that PEI primarily binds via electrostatic interactions to the phosphate backbone, which induces the condensation of the ssDNA. We additionally confirm this finding by conducting molecular dynamics simulations. We expect that the spectral correlations determined here will enable future studies to investigate important gene delivery activities, including how PEI interacts with cellular membranes to facilitate cargo internalization into cells.

探明聚乙烯亚胺（polyethylenimine, PEI）这类聚合物载体如何结合并包装其核酸货物，对于开发更高效、更具成本效益的基因治疗手段至关重要。为了构建DNA结合过程的分子层面图景，我们探究了聚乙烯亚胺的拉曼光谱（Raman spectra）如何反映其局部化学环境。研究发现，位于1400–1500 cm⁻¹区域的强拉曼谱带源自具有显著亚甲基剪式振动与NH弯曲振动特征的分子振动。这些振动对应的拉曼谱带强度变化显著，且同时依赖于局部介电环境与聚合物上仲胺基团参与的氢键相互作用。我们以这些谱带作为光谱标记物，评估了低分子量聚乙烯亚胺与单链DNA（single-stranded DNA, ssDNA）之间的结合作用。拉曼光谱分析表明，聚乙烯亚胺主要通过静电相互作用结合至DNA的磷酸骨架，进而诱导单链DNA发生凝聚。此外，我们通过分子动力学模拟（molecular dynamics simulations）验证了这一发现。我们预期，本研究确定的光谱关联特征将为后续研究提供支撑，以探究重要的基因递送活动，包括聚乙烯亚胺如何与细胞膜相互作用以促进核酸货物向细胞内的内化过程。