生物碱对胃蛋白酶的酶学性质及光谱性质的影响

发布时间：2018-01-28 05:42

  本文关键词： 胃蛋白酶 喜树碱 秋水仙碱 葛根素 紫外光谱 荧光光谱　出处：《四川大学》2005年硕士论文　论文类型：学位论文

【摘要】：胃蛋白酶是动物胃液中主要的蛋白水解酶,具有较强的专一性,催化具有苯丙氨酸,酪氨酸,色氨酸等肽键的断裂,使大分子的蛋白质变为较小的多肽。作为一种重要的蛋白水解酶,胃蛋白酶的酶学性质,底物专一性,抑制作用等都与其它的蛋白酶不同,具用一定的理论研究价值和应用价值。生物碱是一类含氮杂环的碱性天然有机物,广泛存在于植物体内,在医药上有重要的用途。本文基于生物碱的药用价值,借助近年来发展迅速的光谱分析的办法探讨了三种生物碱小分子与胃蛋白酶的相互作用。通过这三种生物碱对胃蛋白酶的动力学性质的影响以及与胃蛋白酶的相互作用的紫外光谱和荧光光谱的研究,对胃蛋白酶在三种小分子影响下的变化及作用机理作了初步推测。 首先对胃蛋白酶的酶学性质进行研究,本实验采用2000药典方法,以血红蛋白作底物,在275nm下以紫外分光光度法测定胃蛋白酶的活力,以每分钟酶水解血红蛋白生成一微摩尔的酪氨酸为一个活力单位。以血红蛋白为底物作酶学性质分析,测得Km值为0.86mmol/L,最适温度为40℃,最适pH2.0。 喜树碱,秋水仙碱,葛根素对胃蛋白酶的动力学影响主要表现在对胃蛋白酶有抑制作用,在不同的底物浓度下分别作三种生物碱对胃蛋白酶的抑制动力学,求得喜树碱ki值为0.63×10~(-3)mol/L,秋水仙碱ki值为0.976×10~(-3)mol/L,葛根素ki值为1.78×10~(-3)mol/L,三种生物碱的抑制剂类型为非竞争性抑制。抑制作用大小依此为:喜树碱秋水仙碱葛根素。 通过对喜树碱,秋水仙碱,葛根素与胃蛋白酶在200-400nm下分别作紫外浓度和时间差谱,发现喜树碱,秋水仙碱与胃蛋白酶在3:1的摩尔浓度比,葛根素与胃蛋白酶在4:1的摩尔浓度比具有最明显的紫外吸收差谱,同时发现处理时间对三种生物碱与胃蛋白酶作用影响不大,说明它们与胃蛋白酶的结合可能是一个瞬时过程。
[Abstract]:Pepsin is the main proteolytic enzyme in animal gastric juice, which has strong specificity and catalyzes the breaking of peptide bonds such as phenylalanine, tyrosine and tryptophan. As an important proteolytic enzyme, pepsin's enzymatic properties, substrate specificity, inhibition and so on are different from other proteases. Alkaloids are a kind of natural alkaloids containing nitrogen heterocycles, which are widely used in plants and have important medical applications. This paper based on the medicinal value of alkaloids. The interaction of three kinds of alkaloids with pepsin was studied by means of the rapidly developing spectral analysis in recent years. The effects of these three alkaloids on the kinetic properties of pepsin and the phase with pepsin were discussed. UV and fluorescence spectra of interaction. The changes and mechanism of pepsin under the influence of three kinds of small molecules were preliminarily speculated. Firstly, the enzymatic properties of pepsin were studied. In this experiment, the activity of pepsin was determined by UV spectrophotometry at 275 nm with hemoglobin as substrate and 2000 pharmacopoeia method. The enzyme hydrolyzed hemoglobin per minute to produce a micromole of tyrosine as a unit of activity, hemoglobin as substrate for enzymatic properties analysis, measured km value of 0.86 mmol / L. The optimum temperature is 40 鈩，

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