香豆素类化合物与乙酰胆碱酯酶的相互作用研究
发布时间：2019-05-15 11:06
【摘要】：阿尔茨海默病是引起中老年人痴呆最常见的疾病。乙酰胆碱酯酶（AChE）抑制剂是当前治疗该疾病的主要手段。目前临床使用的AChE抑制剂存在生物利用率低、活性不是特别高、选择性低、具有不同程度的毒性及副作用等缺点，寻求新的高效低毒的AChE抑制剂是药物化学家们研究的热点之一。香豆素类化合物是具有苯并吡喃环母核的一类天然化合物，研究表明，该类化合物具有一定的AChE抑制活性，具有潜在的药用价值。药物发挥药效的化学本质是药物分子与生物靶分子之间的相互作用，药物药效的高低与其和靶分子之间的相互作用密切相关，因此在分子水平上研究香豆素类化合物与AChE的相互作用，对于探讨该类化合物抑制AChE活性的分子机理及新的AChE抑制活性化合物的筛选和研究都具有重要意义。香豆素类化合物的生物活性与其苯并吡喃环上取代基的种类和位置密切相关，这些取代基团如何影响该类化合物与AChE的相互作用，以及二者之间的体外相互作用与其对酶活性的抑制之间的相关性如何，都是十分值得研究的问题，然而，目前尚未见相关报导。 针对上述问题，本论文选择了结构取代不同的13种香豆素类化合物，通过使用多种光谱方法并结合分子模拟技术研究了上述化合物与AChE之间的相互作用，探讨了该类化合物与AChE相互作用的化学特征和空间结合模式；比较了不同结构取代对该类化合物与AChE相互作用的影响；利用Ellman法对AChE的酶活性进行研究，探讨了不同结构取代对该类化合物对AChE酶活的抑制影响。本研究所获结果对于寻找该类化合物发挥药效的活性基团以及筛选新型的香豆素类AChE抑制剂具有理论的参考意义。具体内容如下： 1香豆素类化合物与AChE相互作用的光谱法研究：利用荧光、紫外-可见吸收光谱法研究了13种香豆素类化合物与AChE的相互作用，探讨了其相互作用的机理，并对相互作用中的结合常数、结合位点数以及二者的结合距离作了计算。由结果可知，该类化合物对AChE的内源荧光具有较强的猝灭作用，猝灭的机制主要为形成复合物的静态猝灭，且能引起AChE构象的变化，疏水作用力和范德华力为其与AChE间相互作用的主要作用力，同时还存在氢键及静电作用力；该类化合物与AChE间的结合距离均小于7nm；不同结构取代化合物相互作用大小的比较表明：香豆素母核4位上羟基的取代以及7位上甲氧基和乙氧基的取代对增强该类化合物与AChE的相互作用具有重要影响。 2香豆素类化合物与AChE相互作用的分子模拟研究：以电鳐乙酰胆碱酯酶的三维空间结构作为模板蛋白，用Autodock4.0软件对AChE与所研究的香豆素类分子进行分子对接，主要对二者相互作用的相互作用能，结合部位及作用力类型进行了判断，结果表明各香豆素化合物均能与AChE的活性位点发生相互作用，且与AChE作用的氨基酸主要为疏水性以及带电荷的氨基酸，在香豆素类分子与AChE形成的复合物中，范德华力及疏水作用力占主导，同时存在氢键与静电作用力，在分子对接研究中对相互作用中作用力类型的判断与荧光方法基本一致；由于香豆素类化合物中香豆素环的存在，其可能与AChE残基产生π-π作用，，π-π共轭也可能是导致AChE荧光猝灭的原因。各个香豆素类化合物由于取代基结构、位置或者个数不同，导致其与AChE相互作用的氨基酸残基种类和个数也不同，与残基形成的氢键个数也有区别，实验结果表明4位上羟基的取代形成的氢键较多。 3香豆素类化合物对AChE活性的改变研究：以碘化硫代乙酰胆碱为底物，利用Ellman法定量地评价了AChE的活性，通过各个化合物对AChE活性的半数抑制浓度的计算来比较各个香豆素类化合物对酶抑制能力的大小。结果表明该类化合物对AChE活性均有一定的抑制作用，香豆素母核4，7位上的取代显示了较好的AChE抑制活性，4，7位上羟基的取代以及羟基取代个数的增加使香豆素类化合物对AChE抑制活性的增强尤为明显。
[Abstract]:Alzheimer's disease is the most common disease in the middle-aged and the elderly. Ethylcholine esterase (AChiE) inhibitors are the main means for the current treatment of the disease. The AChiE inhibitor currently used in clinical use has the disadvantages of low bioavailability, high activity, low selectivity, different degree of toxicity and side effect, and the like, and the novel high-efficiency and low-toxicity AChiE inhibitor is one of the hot spots of the drug chemists. Coumarin compounds are a kind of natural compound with benzo-benzene ring mother nucleus, and the study shows that the compound has certain AChiE inhibitory activity and has potential medicinal value. The chemical nature of the drug effect is the interaction between the drug molecules and the biological target molecules, the drug effect is closely related to the interaction between the drug molecules and the target molecules, It is of great significance to study the molecular mechanism of the compound to inhibit the activity of AChE and the screening and study of the new AChE inhibitory active compound. the biological activity of the coumarin compound is closely related to the species and the position of the substituents on the benzo-homopolar ring, how these substituents affect the interaction of the compound with the achaE, and how the in vitro interaction between the two is related to its inhibition of the enzymatic activity, It is a matter of great value to study, however, no relevant reports have been reported at this time. In the light of the above-mentioned problems, the structure is selected to replace 13 kinds of coumarin compounds, and the interaction between the above-mentioned compounds and the AChE is studied by using a variety of spectral methods and the molecular simulation technology. In this paper, the chemical and spatial-binding modes of the interaction between the compounds and the AChE are discussed. The effect of the substitution of the different structures on the interaction of the compounds with the AChE is compared. The enzyme activity of the AChE is studied by Elman's method. The effect of substitution of different structures on the activity of AChiE in this kind of compound was studied. In response, the results of this study are useful for finding the active groups of these compounds and to screen new type of coumarin AChiE inhibitors. I. Specific contents such as In this paper, the interaction of 13 kinds of coumarin compounds with AChE was studied by fluorescence and ultraviolet-visible absorption spectrometry, and the interaction mechanism was discussed. the combination constant, the combination of the number of bits, and the combined distance of the two The results show that the compounds have a strong quenching effect on the endogenous fluorescence of AChE. The quenching mechanism is mainly to form the static quenching of the complex, and can cause the change of the conformation of the AChE, the hydrophobic force and the van der Waals force are the main factors to interact with the AChE. the interaction distance between the compound and the AChE is less than 7 nm; the ratio of the interaction size of the different structures to the compound It is shown that the substitution of the hydroxyl groups in the 4-position of the coumarin and the substitution of the methoxy and ethoxy groups on the 7-position can be used to enhance the interaction of the compound with the AChE. A molecular simulation study on the interaction of 2-coumarines and AChE was carried out. The three-dimensional space structure of Choline esterase was used as a template protein, and the interaction of AChE with the studied coumarin-based molecules was carried out by Audiock4.0 software. The interaction energy, the binding site and the type of force are determined. The results show that the coumarin compounds can interact with the active sites of the AChiE, and the amino acids that interact with the AChiE are mainly hydrophobic and charged amino acids, which are formed in the coumarin-like molecules and the AChiE. In the complex, the van der Waals force and the hydrophobic force are dominant, and the hydrogen bond and the static force are present at the same time, and the judgment of the force type in the interaction is basically the same as that of the fluorescence method in the molecular docking study; and the coumarins in the coumarin compound The presence of a prime ring, which may be the same as that of the AChE residue, may also be the result of the AChE fluorescence The reason of the quenching is that the number of amino acid residues which can interact with the AChE is different due to the different structure, position or number of the substituents, and the number of hydrogen bonds formed by the residues is different, and the experimental results show that the substitution of the hydroxyl in the 4-position is formed. The changes of the activity of the 3-coumarin compound to the activity of AChE were studied. The activity of AChE was evaluated by Ellman, and the activity of AChE was evaluated by Ellman. The results show that the compound has a certain inhibitory effect on the activity of AChE, the substitution of the 4 and 7 positions of the coumarin shows better AChiE inhibitory activity, the substitution of the hydroxyl groups at the 7-position and the increase of the number of the hydroxyl groups make the coumarin compounds inhibit the AChE.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R969.2
本文编号：2477455
[Abstract]:Alzheimer's disease is the most common disease in the middle-aged and the elderly. Ethylcholine esterase (AChiE) inhibitors are the main means for the current treatment of the disease. The AChiE inhibitor currently used in clinical use has the disadvantages of low bioavailability, high activity, low selectivity, different degree of toxicity and side effect, and the like, and the novel high-efficiency and low-toxicity AChiE inhibitor is one of the hot spots of the drug chemists. Coumarin compounds are a kind of natural compound with benzo-benzene ring mother nucleus, and the study shows that the compound has certain AChiE inhibitory activity and has potential medicinal value. The chemical nature of the drug effect is the interaction between the drug molecules and the biological target molecules, the drug effect is closely related to the interaction between the drug molecules and the target molecules, It is of great significance to study the molecular mechanism of the compound to inhibit the activity of AChE and the screening and study of the new AChE inhibitory active compound. the biological activity of the coumarin compound is closely related to the species and the position of the substituents on the benzo-homopolar ring, how these substituents affect the interaction of the compound with the achaE, and how the in vitro interaction between the two is related to its inhibition of the enzymatic activity, It is a matter of great value to study, however, no relevant reports have been reported at this time. In the light of the above-mentioned problems, the structure is selected to replace 13 kinds of coumarin compounds, and the interaction between the above-mentioned compounds and the AChE is studied by using a variety of spectral methods and the molecular simulation technology. In this paper, the chemical and spatial-binding modes of the interaction between the compounds and the AChE are discussed. The effect of the substitution of the different structures on the interaction of the compounds with the AChE is compared. The enzyme activity of the AChE is studied by Elman's method. The effect of substitution of different structures on the activity of AChiE in this kind of compound was studied. In response, the results of this study are useful for finding the active groups of these compounds and to screen new type of coumarin AChiE inhibitors. I. Specific contents such as In this paper, the interaction of 13 kinds of coumarin compounds with AChE was studied by fluorescence and ultraviolet-visible absorption spectrometry, and the interaction mechanism was discussed. the combination constant, the combination of the number of bits, and the combined distance of the two The results show that the compounds have a strong quenching effect on the endogenous fluorescence of AChE. The quenching mechanism is mainly to form the static quenching of the complex, and can cause the change of the conformation of the AChE, the hydrophobic force and the van der Waals force are the main factors to interact with the AChE. the interaction distance between the compound and the AChE is less than 7 nm; the ratio of the interaction size of the different structures to the compound It is shown that the substitution of the hydroxyl groups in the 4-position of the coumarin and the substitution of the methoxy and ethoxy groups on the 7-position can be used to enhance the interaction of the compound with the AChE. A molecular simulation study on the interaction of 2-coumarines and AChE was carried out. The three-dimensional space structure of Choline esterase was used as a template protein, and the interaction of AChE with the studied coumarin-based molecules was carried out by Audiock4.0 software. The interaction energy, the binding site and the type of force are determined. The results show that the coumarin compounds can interact with the active sites of the AChiE, and the amino acids that interact with the AChiE are mainly hydrophobic and charged amino acids, which are formed in the coumarin-like molecules and the AChiE. In the complex, the van der Waals force and the hydrophobic force are dominant, and the hydrogen bond and the static force are present at the same time, and the judgment of the force type in the interaction is basically the same as that of the fluorescence method in the molecular docking study; and the coumarins in the coumarin compound The presence of a prime ring, which may be the same as that of the AChE residue, may also be the result of the AChE fluorescence The reason of the quenching is that the number of amino acid residues which can interact with the AChE is different due to the different structure, position or number of the substituents, and the number of hydrogen bonds formed by the residues is different, and the experimental results show that the substitution of the hydroxyl in the 4-position is formed. The changes of the activity of the 3-coumarin compound to the activity of AChE were studied. The activity of AChE was evaluated by Ellman, and the activity of AChE was evaluated by Ellman. The results show that the compound has a certain inhibitory effect on the activity of AChE, the substitution of the 4 and 7 positions of the coumarin shows better AChiE inhibitory activity, the substitution of the hydroxyl groups at the 7-position and the increase of the number of the hydroxyl groups make the coumarin compounds inhibit the AChE.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R969.2
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