两种降糖药物与牛血清白蛋白的相互作用及蓝铜蛋白His143性质初步研究
发布时间：2019-04-12 07:07
【摘要】：血清白蛋白是血浆中含量最为丰富的蛋白质,具有贮存转运内源代谢产物和外源药物小分子等重要的生理功能,能与多种内源以及外源分子进行结合,通过形成复合物从而完成血浆的各项生理功能。研究降糖药物与血清白蛋白大分子的相互作用能够在药物代谢动力学、制药学以及新药研发等方提供一定的基础信息。而蓝铜蛋白(RUS)是一类载铜蛋白,是嗜酸氧化亚铁硫杆菌中铁呼吸电子传递的重要组分,研究该蛋白的性质对铜蛋白家族中的其他蛋白如生物血浆铜蓝蛋白等性质结构提供一定的依据。本论文采用荧光光谱技术和紫外-可见光技术为研究手段,研究了两种降糖药物(盐酸吡格列酮(PIO)和盐酸二甲双胍(MET))与牛血清白蛋白(BSA)的相互作用,探究了这两种降糖药物与BSA的作用方式,为降糖药的研发以及更好的指导用药提供依据。同时采用光谱法和分子结构模拟技术研究RUS His143在电子传递中的作用,并通过对His143定点突变进一步验证。主要工作内容如下:1.光谱法研究PIO与BSA的相互作用。采用荧光光谱法和紫外-可见光谱法,发现PIO对BSA荧光有明显猝灭作用,且其猝灭方式为动态猝灭;进一步测定了PIO与BSA的结合常数K、结合位点数及ΔG、ΔH、ΔS等热力学常数;并根据三个热力学常数确定PIO与BSA之间的相互作用为疏水作用力。2.光谱法研究MET与BSA的相互作用。采用荧光光谱法等方法初步探究MET对BSA荧光的猝灭作用,得到其猝灭方式为动态猝灭;根据测定的MET与BSA的结合常数K和结合位点数及热力学常数,确定MET与BSA之间的相互作用为氢键和范德华力,且通过分子对接方法进一步验证该结论。3.光谱法和分子模拟技术研究RUS His143的性质。通过对His143位点的定点突变,聚丙烯酰胺凝胶电泳,光谱学数据初步确定His143位点是RUS与外界电子传递的重要位点,并结合分子结构模拟结果显示His143位点的咪唑基团形成疏水屏障从而使铜离子不会暴露于溶液环境,进一步说明RUS His143在电子传递中的重要性。
[Abstract]:Serum albumin is the most abundant protein in plasma. It has important physiological functions such as storing and transporting endogenous metabolites and small molecules of exogenous drugs. It can combine with many kinds of endogenous and exogenous molecules. The physiological functions of the plasma are accomplished by the formation of a complex. The study of the interaction between hypoglycemic drugs and serum albumin can provide some basic information in pharmacokinetic, pharmaceutical and new drug development. Blue copper protein (RUS) is a kind of copper-carrying protein, which is an important component of iron respiratory electron transport in thiobacillus ferrooxidans. The study of the properties of the protein provides a basis for the structure of other proteins in the copper protein family, such as plasma ceruloplasmin. In this paper, the interaction of pioglitazone hydrochloride (PIO) and metformin hydrochloride (MET) with bovine serum albumin (BSA) was studied by fluorescence spectroscopy and ultraviolet-visible light (UV-vis) techniques, and the interaction of pioglitazone hydrochloride and metformin hydrochloride with bovine serum albumin (BSA) was studied. The effects of these two hypoglycemic drugs on BSA were explored, which provided the basis for the research and development of hypoglycemic drugs and better guidance for drug use. At the same time, the role of RUS His143 in electron transfer was studied by spectroscopic and molecular structure simulation techniques, and further verified by site-directed mutation of His143. The main contents of the work are as follows: 1. The interaction between PIO and BSA was studied by spectroscopic method. By means of fluorescence and UV-vis spectroscopy, it was found that PIO had obvious quenching effect on BSA fluorescence, and its quenching mode was dynamic quenching. The binding constants K, binding site number and 螖 G, 螖 H, 螖 S of PIO and BSA were further determined, and the interaction between PIO and BSA was determined to be hydrophobic force according to the three thermodynamic constants. The interaction between MET and BSA was studied by spectroscopic method. The quenching effect of MET on BSA fluorescence was studied by fluorescence spectroscopy and other methods. The quenching mode was dynamic quenching. According to the binding constants K and the number of binding sites and thermodynamic constants of MET and BSA, the interaction between MET and BSA is determined as hydrogen bond and van der Waals force, and the conclusion is further verified by molecular docking method. Spectroscopic and molecular simulation techniques were used to study the properties of RUS His143. By site-directed mutation of His143 site, polyacrylamide gel electrophoresis (page) and spectroscopic data, the His143 site was determined to be an important site for electron transfer between RUS and the outside world. The results of molecular structure simulation show that the imidazole group at His143 site forms a hydrophobic barrier so that copper ions will not be exposed to the solution environment, which further illustrates the importance of RUS His143 in electron transport.
【学位授予单位】：湖南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R96
本文编号：2456813
[Abstract]:Serum albumin is the most abundant protein in plasma. It has important physiological functions such as storing and transporting endogenous metabolites and small molecules of exogenous drugs. It can combine with many kinds of endogenous and exogenous molecules. The physiological functions of the plasma are accomplished by the formation of a complex. The study of the interaction between hypoglycemic drugs and serum albumin can provide some basic information in pharmacokinetic, pharmaceutical and new drug development. Blue copper protein (RUS) is a kind of copper-carrying protein, which is an important component of iron respiratory electron transport in thiobacillus ferrooxidans. The study of the properties of the protein provides a basis for the structure of other proteins in the copper protein family, such as plasma ceruloplasmin. In this paper, the interaction of pioglitazone hydrochloride (PIO) and metformin hydrochloride (MET) with bovine serum albumin (BSA) was studied by fluorescence spectroscopy and ultraviolet-visible light (UV-vis) techniques, and the interaction of pioglitazone hydrochloride and metformin hydrochloride with bovine serum albumin (BSA) was studied. The effects of these two hypoglycemic drugs on BSA were explored, which provided the basis for the research and development of hypoglycemic drugs and better guidance for drug use. At the same time, the role of RUS His143 in electron transfer was studied by spectroscopic and molecular structure simulation techniques, and further verified by site-directed mutation of His143. The main contents of the work are as follows: 1. The interaction between PIO and BSA was studied by spectroscopic method. By means of fluorescence and UV-vis spectroscopy, it was found that PIO had obvious quenching effect on BSA fluorescence, and its quenching mode was dynamic quenching. The binding constants K, binding site number and 螖 G, 螖 H, 螖 S of PIO and BSA were further determined, and the interaction between PIO and BSA was determined to be hydrophobic force according to the three thermodynamic constants. The interaction between MET and BSA was studied by spectroscopic method. The quenching effect of MET on BSA fluorescence was studied by fluorescence spectroscopy and other methods. The quenching mode was dynamic quenching. According to the binding constants K and the number of binding sites and thermodynamic constants of MET and BSA, the interaction between MET and BSA is determined as hydrogen bond and van der Waals force, and the conclusion is further verified by molecular docking method. Spectroscopic and molecular simulation techniques were used to study the properties of RUS His143. By site-directed mutation of His143 site, polyacrylamide gel electrophoresis (page) and spectroscopic data, the His143 site was determined to be an important site for electron transfer between RUS and the outside world. The results of molecular structure simulation show that the imidazole group at His143 site forms a hydrophobic barrier so that copper ions will not be exposed to the solution environment, which further illustrates the importance of RUS His143 in electron transport.
【学位授予单位】：湖南大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R96
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