細(xì)胞色素P450酶介導(dǎo)的芳香族化合物代謝機(jī)理的研究
發(fā)布時間：2018-11-29 13:08
【摘要】：芳香族化合物(Aromatic Compounds)的來源與分布都非常廣泛,是環(huán)境中典型的有機(jī)污染物。該類化合物的分子結(jié)構(gòu)中一般都帶有苯環(huán),當(dāng)其進(jìn)入生物體后極易引發(fā)致畸、致癌、致突變效應(yīng),對人類的健康產(chǎn)生巨大的威脅。細(xì)胞色素P450酶(Cytochrome P450)作為自然界中的萬能氧化酶對進(jìn)入生物體的外源性化合物尤其是一些間接致癌物的代謝轉(zhuǎn)化(包括代謝解毒和代謝活化)中起著至關(guān)重要的作用。這類化合物進(jìn)入生物體后,經(jīng)細(xì)胞色素P450酶的氧化激活生成極易與體內(nèi)的DNA、蛋白質(zhì)等生物大分子上的親核位點發(fā)生共價結(jié)合的親電分子,引起腫瘤,誘發(fā)癌癥。因此,深入了解這類化合物在P450酶介導(dǎo)下的代謝路徑具有極其重要的意義,這有助于診斷環(huán)境污染的狀況和程度,并為早期環(huán)境預(yù)警提供理論支撐。但由于常規(guī)的實驗手段存在諸多限制,例如,P450酶的活性單體CpdⅠ(CompoundⅠ)、Cpd0(Compound 0)等在反應(yīng)過程中轉(zhuǎn)瞬即逝,極其難以捕捉,給化學(xué)反應(yīng)動力學(xué)的研究帶來極大的困難。因此我們采用密度泛函理論(Density Functional Theory,DFT)計算的方法來對這些反應(yīng)進(jìn)行研究。主要分為以下三部分內(nèi)容:(1)本位取代反應(yīng)(ipso-Substitution)是P450酶代謝對位取代苯酚類化合物的一種重要途徑,卻鮮有用量子化學(xué)計算的方法來研究其內(nèi)在機(jī)制的報道。我們以雙酚A為模型化合物對其在P450酶的CpdⅠ介導(dǎo)下的本位取代反應(yīng)路徑與氧加成路徑進(jìn)行計算,比較后發(fā)現(xiàn)本位取代反應(yīng)路徑為優(yōu)勢反應(yīng)路徑,即:P450酶的CpdⅠ活性中心的氧將酚上的氫提取后反彈到對位或鄰位生成相應(yīng)產(chǎn)物,該路徑發(fā)生在低自旋二重態(tài)下,速率決定步驟為氫原子的提取。同時我們研究了一系列對位不同取代基苯酚發(fā)現(xiàn)本位取代反應(yīng)的過渡態(tài)能壘與取代基效應(yīng)密切相關(guān),隨對位取代基特性常數(shù)σp的增大而增大。(2)苯并(a)花(Benzo[a]pyrene)作為一種典型的P450酶底物,具有極強(qiáng)的致癌效應(yīng)。本研究從反應(yīng)動力學(xué)及熱力學(xué)方面對P450酶的CpdⅠ環(huán)氧化苯并(a)芘各活性位點的路徑進(jìn)行比較,結(jié)合前人關(guān)于離域能的研究進(jìn)一步解釋了該反應(yīng)中出現(xiàn)的區(qū)域選擇性現(xiàn)象。并對該反應(yīng)的主產(chǎn)物之一——9,10-環(huán)氧化苯并芘的形成路徑進(jìn)行分析,得出該反應(yīng)主要發(fā)生在低自旋二重態(tài)的結(jié)論。(3)芳香胺類化合物(Aromatic Amine)可以在P450酶的CpdⅠ作用下代謝活化生成氮正陽離子自由基,與體內(nèi)的親核分子發(fā)生共價結(jié)合誘發(fā)癌癥。本研究通過DFT方法計算了包括普通芳香胺與氮雜環(huán)芳香胺在內(nèi)的28中芳香胺類化合物氫在P450酶的CpdⅠ介導(dǎo)下的兼并雙態(tài)下氫提取反應(yīng)的平均活化能壘AE*,發(fā)現(xiàn)該能壘△E**與N-H鍵的解離能(Bond Dissociation Energy,BDEN-H)之間存在著良好的線性相關(guān)。通過BDEN-H與AE*建立的預(yù)測模型(簡單芳香胺:R2=0.92;氮雜環(huán)芳香胺:R2=0.95)具有良好的穩(wěn)健性和預(yù)測能力。這為快速有效預(yù)測此類化合物的代謝活性甚至潛在的致癌性風(fēng)險提供了依據(jù)。
[Abstract]:The source and distribution of aromatic compounds are very broad and are typical organic pollutants in the environment. The molecular structure of the compound is generally provided with a benzene ring, and when the benzene ring enters an organism, a teratogenic, carcinogenic and mutagenic effect can be easily induced, and a great threat to the health of the human is generated. Cytochrome P450 (Cytochrome P450), as a universal oxidase in nature, plays a critical role in the metabolic transformation of exogenous compounds, especially some indirect carcinogens, into organisms, including metabolic detoxification and metabolic activation. after the compound enters the organism, the cell pigment P450 enzyme is oxidized and activated to generate the electrophilic molecule which is very easy to be covalently bound to the nucleon site on the biological macromolecule such as the DNA and the protein in the body, and causes the tumor and the cancer to be induced. Therefore, it is of great significance to understand the metabolic pathway of these compounds under the P450 enzyme, which can help to diagnose the condition and extent of environmental pollution and provide theoretical support for early environmental early warning. However, due to the limitations of conventional experimental means, for example, the activity monomer Cpd I (Sound I), Cpd0 (Sound 0), and the like of the P450 enzyme are transient in the course of the reaction, and it is extremely difficult to capture, which brings great difficulty to the study of the chemical reaction kinetics. So we use the density functional theory (DFT) to study these reactions. It is mainly divided into the following three parts: (1) the standard substitution reaction (ipso-subregulation) is an important way for P450 enzyme to metabolize the para-substituted benzene phenolic compound, but few of the methods of quantum chemistry calculation are used to study the report of the mechanism. We use diphenol A as a model compound to calculate the standard substitution reaction path and the oxygen addition path mediated by the Cpd I of the P450 enzyme, and then find that the standard substitution reaction path is the advantage reaction path, namely: The oxygen of the active center of the Cpd I of the P450 enzyme returns to the para-position or the adjacent position to generate a corresponding product after the oxygen on the phenol is extracted, and the path takes place in the low-spin double state, and the rate determination step is the extraction of the hydrogen atom. At the same time, we have studied the transition state energy base of a series of p-substituted-based phenol-based substitution reaction, which is closely related to the substitution-based effect, and increases with the increase of the p-substitution-based property constant (p). (2) Benzo[a] pyrene, as a typical P450 enzyme substrate, has a very strong carcinogenic effect. In this study, from the reaction kinetics and the thermodynamic aspect, the Cpd I-ring of the P450 enzyme was compared with the path of each active site, and the regional selectivity in the reaction was further explained in the light of the previous study on the off-domain energy. The formation path of one of the main products of the reaction is analyzed, and the conclusion that the reaction is mainly in the low-spin double state is obtained. (3) The aromatic amine compound can be metabolically activated under the action of the Cpd I of the P450 enzyme to generate the positive cationic radical of the nitrogen, and is covalently bound to the nucleophile in the body to induce the cancer. In this study, the average activation energy base AE * of the hydrogen extraction reaction of the hydrogen in 28 of the 28 aromatic amine compounds, including the common aromatic amine and the nitrogen heterocyclic aromatic amine, was calculated by the DFT method, and the release energy (Bond Disassociation Energy) of the energy barrier E ** and the N-H bond was found. There is a good linear correlation between BDEN-H. The prediction model established by BDEN-H and AE * (simple aromatic amine: R2 = 0.92; N-heterocyclic aromatic amine: R2 = 0.95) has good robustness and prediction ability. This provides a basis for rapid and effective prediction of the metabolic activity of such compounds and even the potential carcinogenicity risk.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：X592
本文編號：2365080
[Abstract]:The source and distribution of aromatic compounds are very broad and are typical organic pollutants in the environment. The molecular structure of the compound is generally provided with a benzene ring, and when the benzene ring enters an organism, a teratogenic, carcinogenic and mutagenic effect can be easily induced, and a great threat to the health of the human is generated. Cytochrome P450 (Cytochrome P450), as a universal oxidase in nature, plays a critical role in the metabolic transformation of exogenous compounds, especially some indirect carcinogens, into organisms, including metabolic detoxification and metabolic activation. after the compound enters the organism, the cell pigment P450 enzyme is oxidized and activated to generate the electrophilic molecule which is very easy to be covalently bound to the nucleon site on the biological macromolecule such as the DNA and the protein in the body, and causes the tumor and the cancer to be induced. Therefore, it is of great significance to understand the metabolic pathway of these compounds under the P450 enzyme, which can help to diagnose the condition and extent of environmental pollution and provide theoretical support for early environmental early warning. However, due to the limitations of conventional experimental means, for example, the activity monomer Cpd I (Sound I), Cpd0 (Sound 0), and the like of the P450 enzyme are transient in the course of the reaction, and it is extremely difficult to capture, which brings great difficulty to the study of the chemical reaction kinetics. So we use the density functional theory (DFT) to study these reactions. It is mainly divided into the following three parts: (1) the standard substitution reaction (ipso-subregulation) is an important way for P450 enzyme to metabolize the para-substituted benzene phenolic compound, but few of the methods of quantum chemistry calculation are used to study the report of the mechanism. We use diphenol A as a model compound to calculate the standard substitution reaction path and the oxygen addition path mediated by the Cpd I of the P450 enzyme, and then find that the standard substitution reaction path is the advantage reaction path, namely: The oxygen of the active center of the Cpd I of the P450 enzyme returns to the para-position or the adjacent position to generate a corresponding product after the oxygen on the phenol is extracted, and the path takes place in the low-spin double state, and the rate determination step is the extraction of the hydrogen atom. At the same time, we have studied the transition state energy base of a series of p-substituted-based phenol-based substitution reaction, which is closely related to the substitution-based effect, and increases with the increase of the p-substitution-based property constant (p). (2) Benzo[a] pyrene, as a typical P450 enzyme substrate, has a very strong carcinogenic effect. In this study, from the reaction kinetics and the thermodynamic aspect, the Cpd I-ring of the P450 enzyme was compared with the path of each active site, and the regional selectivity in the reaction was further explained in the light of the previous study on the off-domain energy. The formation path of one of the main products of the reaction is analyzed, and the conclusion that the reaction is mainly in the low-spin double state is obtained. (3) The aromatic amine compound can be metabolically activated under the action of the Cpd I of the P450 enzyme to generate the positive cationic radical of the nitrogen, and is covalently bound to the nucleophile in the body to induce the cancer. In this study, the average activation energy base AE * of the hydrogen extraction reaction of the hydrogen in 28 of the 28 aromatic amine compounds, including the common aromatic amine and the nitrogen heterocyclic aromatic amine, was calculated by the DFT method, and the release energy (Bond Disassociation Energy) of the energy barrier E ** and the N-H bond was found. There is a good linear correlation between BDEN-H. The prediction model established by BDEN-H and AE * (simple aromatic amine: R2 = 0.92; N-heterocyclic aromatic amine: R2 = 0.95) has good robustness and prediction ability. This provides a basis for rapid and effective prediction of the metabolic activity of such compounds and even the potential carcinogenicity risk.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：X592
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