光谱法研究三种头孢类药物与胰蛋白酶、牛转铁蛋白之间的相互作用

发布时间：2018-05-30 14:11

本文选题：光谱法 + 头孢类药物　；参考：《河北大学》2017年硕士论文

【摘要】：近年来,人类疾病的发生率逐年增加,各种药物在生活中也起到了不容忽视的作用,为了更好的了解药物的药理、药效和靶向运输,药物与蛋白相互作用的研究课题已成为人们关注的重点。本文以胰蛋白酶和牛转铁蛋白为载体蛋白,用多种方法研究了硫酸头孢匹罗、头孢哌酮钠、头孢尼西钠与蛋白的相互作用。研究内容分为五部分:第一章:综述了几种研究蛋白与小分子药物相互作用的常用方法,并对胰蛋白酶和转铁蛋白进行了简单的概述。对文章的立题思路、研究内容以及药物与蛋白相互作用的展望进行了简单的介绍。第二章:用多种光谱法和分子对接法研究了胰蛋白酶和硫酸头孢匹罗之间的相互作用。结果表明,硫酸头孢匹罗使胰蛋白酶发生荧光猝灭,猝灭机理为静态猝灭,结合位点数接近于1。猝灭曲线表明色氨酸残基和酪氨酸残基都参加了反应。同步荧光光谱法进一步表明,二者主要的结合位点更接近于色氨酸残基。对热力学参数的计算表明,二者之间主要作用力为静电引力,分子对接表明二者之间的作用力为静电引力,其中还有氢键作用。胰蛋白酶与硫酸头孢匹罗之间的距离小于7 nm,表明从胰蛋白酶到硫酸头孢匹罗的能量转移为非辐射能量转移。荧光光谱法对Hill系数的研究表明,硫酸头孢匹罗与胰蛋白酶之间存在负协同作用。第三章:利用荧光光谱法,同步荧光光谱法和紫外吸收光谱法研究了硫酸头孢匹罗和牛转铁蛋白之间的相互作用。测定了二者之间的猝灭常数、结合常数、结合位点等。通过对热力学参数的计算,确定了硫酸头孢匹罗和牛转铁蛋白之间的主要作用力类型以静电引力为主。通过对Hill系数的计算,得出了硫酸头孢匹罗和牛转铁蛋白之间的协同作用为负协同作用。第四章:通过荧光猝灭法和同步荧光光谱法对头孢哌酮钠与牛转铁蛋白的相互作用机理进行了研究。结果表明,两种方法用相同的公式计算得到的二者的猝灭机理、结合位点,相互作用力以及协同作用基本一致。并且通过对Δλ=60 nm和λex=295 nm的数据进行比较,得出同步荧光光谱法较之于传统荧光光谱法有更高的灵敏度和准确度。因此,同步荧光光谱法极大地丰富了小分子配体与蛋白之间相互作用机理的研究方法。第五章:实验在pH=7.4缓冲溶液条件下,研究了不同温度下头孢尼西钠与牛转铁蛋白的相互作用。结果表明,牛转铁蛋白-头孢尼西钠体系中,主要的作用方式为生成了新的复合物的静态猝灭,二者的结合位点数为1。由热力学参数可知头孢尼西钠与牛转铁蛋白主要通过静电引力结合。紫外吸收光谱法和同步荧光光谱法都证明了头孢尼西钠使得牛转铁蛋白氨基酸极性增大,疏水性减小。
[Abstract]:In recent years, the incidence of human diseases has increased year by year, and various drugs have played an important role in life. In order to better understand the pharmacology, efficacy and targeted transportation of drugs, The study of drug-protein interaction has become the focus of attention. Using trypsin and bovine transferrin as carrier proteins, the interaction of cefpirosine sulfate, cefoperazone sodium and cefnesian sodium with protein was studied by various methods. The research contents are divided into five parts: chapter 1: the common methods of studying the interaction between protein and small molecule drugs are reviewed, and the trypsin and transferrin are briefly summarized. The idea, research content and prospect of drug-protein interaction are briefly introduced in this paper. In chapter 2, the interaction between trypsin and cefpirome sulfate was studied by multiple spectral and molecular docking methods. The results showed that cefpirosine sulfate caused fluorescence quenching of trypsin, the quenching mechanism was static quenching, and the binding site number was close to 1. The quenching curves showed that both tryptophan residues and tyrosine residues participated in the reaction. Synchronous fluorescence spectroscopy further showed that the binding sites of the two were closer to tryptophan residues. The calculation of thermodynamic parameters shows that the main force between them is electrostatic force, and the molecular docking shows that the force between them is electrostatic force, among which there is hydrogen bond. The distance between trypsin and cefpirosine sulfate is less than 7 nm, which indicates that the energy transfer from trypsin to cefpirome sulfate is non-radiation energy transfer. The study of Hill coefficient by fluorescence spectroscopy showed that there was a negative synergistic effect between cefopirol sulfate and trypsin. In chapter 3, the interaction between cefpirosine sulfate and bovine transferrin was studied by fluorescence spectroscopy, synchronous fluorescence spectroscopy and ultraviolet absorption spectrometry. The quenching constants and binding sites were determined. Based on the calculation of thermodynamic parameters, the main force between cefopiro sulfate and bovine transferrin was determined to be electrostatic force. By calculating the Hill coefficient, the synergism between cefopirol sulfate and bovine transferrin was found to be negative. Chapter 4: the interaction mechanism between cefoperazone sodium and bovine transferrin was studied by fluorescence quenching and synchronous fluorescence spectrometry. The results show that the quenching mechanism, binding site, interaction force and synergism between the two methods calculated by the same formula are basically the same. By comparing the data of 螖位 ~ (60) nm and 位 ex=295 nm, it is concluded that the synchronous fluorescence spectrum method has higher sensitivity and accuracy than the traditional fluorescence spectrum method. Therefore, synchronous fluorescence spectroscopy has greatly enriched the mechanism of interaction between small molecular ligands and proteins. Chapter 5: the interaction between cefnesian sodium and bovine transferrin at different temperatures in pH=7.4 buffer solution was studied. The results showed that the main action mode of bovine transferin-cefnesian sodium system was the static quenching of the formation of new complexes, the binding site of which was 1. Thermodynamic parameters show that cefnesian sodium binds to bovine transferrin mainly by electrostatic force. Both UV absorption and synchronous fluorescence spectra showed that cefnesian sodium increased amino acid polarity and decreased hydrophobicity of bovine transferrin.
【学位授予单位】：河北大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R96;O657.3

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