磷酰化黄酮与钙结合型蛋白的弱相互作用研究
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摘要
细胞功能多是由生物分子之间的弱相互作用引起的，它们包括：酶与底物之间，蛋白质与配体之间，蛋白质与蛋白质之间以及抗体与抗原。在生物体中，蛋白质是必不可少的生命物质，在生命的运动和发展延续中起着重要的作用。有关蛋白质构象、组成结构、以及和小分子相互作用的机理，是目前生命科学、化学和临床医学中共同关注和感兴趣的课题。近年来，由于医药学和生命科学的发展，研究小分子，尤其是具有药效的小分子与蛋白质的相互作用机理过程，对药代动力学认识和开发新药有极其重要的意义。自从电喷雾质谱问世以来，它已经成为一种检测气相中跟大分子相关的弱相互作用的有力工具。
许多事实表明磷酰胺及其酯类衍生物大多具有广泛的生物活性，在许多生理过程中起着非常重要的作用。黄酮类化合物与大分子的作用比较弱，作为有机磷小分子化合物，其磷酰酯类衍生物大多具有广泛的生物活性。本文将经改造的并较简化的Atheron-Todd反应扩大到黄酮和异黄酮的磷酰化结构改造上，选择了7-羟基黄酮，5，7-二羟基黄酮，成功合成了一系列新的磷酰化黄酮。
本文以现代生物质谱ESI，开展了4种磷酰化黄酮和黄酮的与α-乳白蛋白蛋白发生弱相互作用的研究，ESI质谱法的结果显示了4种磷酰化黄酮产物的确能够和α-乳白蛋白发生弱相互作用，它们跟α-乳白蛋白不同程度的结合，证明它们与α-乳白蛋白之间有着有较强的亲和力。而我们在相同的ESI质谱条件(甚至锥孔电压更低)下检测黄酮本身与α-乳白蛋白的弱相互作用，却未发现复合物的存在。确立磷酰基在分子与蛋白发生相互作用时的关键性的作用。提高锥孔电压其实就是提高了复合物的内能，使其变的不稳定。本文将取样孔的电压(锥孔电压)设置很高(接近120V)，而仍能观察到复合物的存在，这就说明，α-乳白蛋白与4种磷酰化黄酮形成的复合物的结合能相对较强。此事实更进一步证明了经过磷酰化修饰的黄酮的生物活性得到提高，也因而更有可能成为一种潜在的抑制剂。
除了α-乳白蛋白，本文还用荧光法研究了磷酰化黄酮与另外一种钙离子结合性蛋白——钙调素蛋白之间的弱相互作用。本文参考文献从牛脑、牛心中分别提
Cellular functions are often triggered by weak non-covalent enzyme-substrate, protein-ligand, protein-protein or antibody-antigen interactions. In organisms, proteins are vital substances, being very important during the course of movement and development of life. The mechanisms about proteins' structure and constitutes and the interaction with small molecules are topics interested by life scientists and chemists and clinic doctors. Recently, duing to the development of life scinence and medicine scinence, studies on the interaction between small molecules, especially medicative small ones and proteins have good meanings to the empolder of new drugs. Since its introduction, ESI-MS has served a powerful tool in providing evidence in support of the existence of non-covalently associated marcromolecular complexes in the gas phase.
It is an established fact that esters of phosphoric acid have wide bio-activities and play a vital role in many biological processes.In the study described in this thesis, aminophenols and dihydroxybenzenes were first phosphorylated by modification of the classical Atheron-Todd procedure in our primary stage; Then the mechanism of the reaction between salicylic acid and dialkyl phosphite was studied by electrospray ionozation mass spectrometry and ~(31)P NMR; 4-aminoantipine was further phosphorylated by modification of the classical Atheron-Todd procedure; finally, we have been able to enlarge the scope of the Atheron-Todd reaction to flavone.
Electrospray ionization results showed that the phosphorylated flavonoids could form non-covalent complexes with many proteins such as lysozyme, myoglobin, bovine insulin and cytochrome c, while non-covalent complexes were not detected with the mixed solution of the chrysin and proteins. The phosphorylated flavonoids possess relatively stronger affinities and form non-covalent complexes with the proteins more easily than the unphosphorylated compounds.
α-Lactalbumin is a protein deserved attention according to its composing of amino acids. The interaction between α-lactalbumin and dithyl flavon-7-yl phosphate
许多事实表明磷酰胺及其酯类衍生物大多具有广泛的生物活性，在许多生理过程中起着非常重要的作用。黄酮类化合物与大分子的作用比较弱，作为有机磷小分子化合物，其磷酰酯类衍生物大多具有广泛的生物活性。本文将经改造的并较简化的Atheron-Todd反应扩大到黄酮和异黄酮的磷酰化结构改造上，选择了7-羟基黄酮，5，7-二羟基黄酮，成功合成了一系列新的磷酰化黄酮。
本文以现代生物质谱ESI，开展了4种磷酰化黄酮和黄酮的与α-乳白蛋白蛋白发生弱相互作用的研究，ESI质谱法的结果显示了4种磷酰化黄酮产物的确能够和α-乳白蛋白发生弱相互作用，它们跟α-乳白蛋白不同程度的结合，证明它们与α-乳白蛋白之间有着有较强的亲和力。而我们在相同的ESI质谱条件(甚至锥孔电压更低)下检测黄酮本身与α-乳白蛋白的弱相互作用，却未发现复合物的存在。确立磷酰基在分子与蛋白发生相互作用时的关键性的作用。提高锥孔电压其实就是提高了复合物的内能，使其变的不稳定。本文将取样孔的电压(锥孔电压)设置很高(接近120V)，而仍能观察到复合物的存在，这就说明，α-乳白蛋白与4种磷酰化黄酮形成的复合物的结合能相对较强。此事实更进一步证明了经过磷酰化修饰的黄酮的生物活性得到提高，也因而更有可能成为一种潜在的抑制剂。
除了α-乳白蛋白，本文还用荧光法研究了磷酰化黄酮与另外一种钙离子结合性蛋白——钙调素蛋白之间的弱相互作用。本文参考文献从牛脑、牛心中分别提
Cellular functions are often triggered by weak non-covalent enzyme-substrate, protein-ligand, protein-protein or antibody-antigen interactions. In organisms, proteins are vital substances, being very important during the course of movement and development of life. The mechanisms about proteins' structure and constitutes and the interaction with small molecules are topics interested by life scientists and chemists and clinic doctors. Recently, duing to the development of life scinence and medicine scinence, studies on the interaction between small molecules, especially medicative small ones and proteins have good meanings to the empolder of new drugs. Since its introduction, ESI-MS has served a powerful tool in providing evidence in support of the existence of non-covalently associated marcromolecular complexes in the gas phase.
It is an established fact that esters of phosphoric acid have wide bio-activities and play a vital role in many biological processes.In the study described in this thesis, aminophenols and dihydroxybenzenes were first phosphorylated by modification of the classical Atheron-Todd procedure in our primary stage; Then the mechanism of the reaction between salicylic acid and dialkyl phosphite was studied by electrospray ionozation mass spectrometry and ~(31)P NMR; 4-aminoantipine was further phosphorylated by modification of the classical Atheron-Todd procedure; finally, we have been able to enlarge the scope of the Atheron-Todd reaction to flavone.
Electrospray ionization results showed that the phosphorylated flavonoids could form non-covalent complexes with many proteins such as lysozyme, myoglobin, bovine insulin and cytochrome c, while non-covalent complexes were not detected with the mixed solution of the chrysin and proteins. The phosphorylated flavonoids possess relatively stronger affinities and form non-covalent complexes with the proteins more easily than the unphosphorylated compounds.
α-Lactalbumin is a protein deserved attention according to its composing of amino acids. The interaction between α-lactalbumin and dithyl flavon-7-yl phosphate
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