光谱法研究硫酸酯化壳聚糖与DNA的作用机理

Study on the Binding Mode of Sulfated Chitosans with Nucleic Acids Using Spectral Analysis

以荧光光谱法、吸收光谱法、碱变性曲线、离子强度和荧光猝灭等方法研究了两种硫酸酯化壳聚糖与DNA的相互作用机理。结果表明,硫酸酯化壳聚糖与荧光探针DNA/EB的作用存在嵌插和静电作用两种模式。高取代度硫酸酯化壳聚糖(CT-H)与低取代度硫酸酯化壳聚糖(CT-L)存在下,DNA的紫外吸收光谱产生不同的增色效应和减色效应;碱变性pH增大、发光体系稳定性增加;金属阳离子Mg2+可与DNA的磷酸基团产生弱静电相互作用。极微量硫酸酯化壳聚糖存在下,荧光被有效地猝灭,证明在EB、CT-H或CT-L和DNA之间产生了强的竞争键合作用,表明不同硫酸酯化壳聚糖是一种有希望的基因治疗靶向分子。

The interaction of two kinds of sulfated chitosans with DNA was investigated using fluorescence spectra, absorption spectra, alkaline denaturation curves, ionic strength, fluorescence quenching etc. The results showed that there were two modes of the interaction between sulfated chitosans and DNA/ethidium bromide（EB） ： intercalative and electrostatic binding. Hyperchromic and bathochromic effects in the absorption spectra were observed when sulfated chitosans with a low degree of substitution （CT-L） or a high degree of substitution （CT-L） were bound to herring sperm DNA. The stability of DNA/EB was increased with the increase in alkaline denaturation pH in the presence of CT-H and CT-L. Meanwhile, a weak interaction of cationic Mg^2＋ and phosphate groups of DNA by electrostatic force binding was observed. The fluorescence was efficiently quenched in the presence of a micro-amounts of sulfated chitosans, which confirmed that there was a strong competitive binding interaction among EB, CT-H or CT-L and DNA. All of these demonstrated that different sulfated chitosans can be used as a type of hopeful target-pointed molecule of gene therapy.