人α1微球蛋白质的结构和功能研究
发布时间：2018-08-11 09:43
【摘要】：背景脂质运载蛋白是一种拥有30-50个成员的蛋白质超家族，有着保守的三维结构，但却发挥着一系列不同的生物学功能。作为脂质运载蛋白质超家族的成员之一， α1-微球蛋白质（α1-microglobulin, α1M）在进化上高度保守且分布广泛提示该蛋白质具有重要的生理功能，目前有研究发现其能结合和降解血红素，具有还原酶活性及抗氧化性及免疫调节作用。但其功能活性位点以及具体的功能机制仍然不清楚。 目的本研究将解析人α1M蛋白质的三维结构并基于其结构研究其生化和生理功能，从而了解其具体的功能机制并为基于α1M蛋白质三维结构的抗炎、抗氧化及免疫调节药物的研究开发打下基础。 方法通过克隆、表达纯化等技术获得足够纯度的人α1M蛋白质，然后培养蛋白质晶体，通过X衍射技术获得晶体衍射数据随之解析人α1M蛋白质三维结构；基于其三维结构，利用定点突变，光谱分析等技术研究α1M蛋白质的功能机制。 结果通过分子克隆、表达纯化等相关技术成功获得了纯度达95%的人α1M蛋白质；分子筛（Superdex75）纯化结果显示α1M在溶液中以单体和二聚体两种聚集状态存在；通过蛋白质晶体的筛选及优化成功获得了适于X射线衍射的高质量单晶；收集到了分辨率为2.0常规衍射数据及分辨率为3.0单波长反常散射衍射(single-wavelengthanomalous dispersion，，SAD)数据，成功的解析了α1M蛋白质的三维结构；利用ABTS法进行检测，结果表明本研究获得重组的α1M具有抗氧化能力，具备还原酶活性。
[Abstract]:Background Lipid transport protein is a protein superfamily with 30-50 members, which has a conserved three-dimensional structure, but plays a series of different biological functions. As one of the members of lipid transport protein superfamily, 伪 1-microglobulin (伪 1m) is highly conserved and widely distributed in evolution. It has been found that 伪 1-microglobulin (伪 1m) can bind and degrade heme. It has reductase activity, antioxidant activity and immunomodulatory effect. However, its functional active sites and specific functional mechanisms are still unclear. Objective in this study, the three-dimensional structure of human 伪 1m protein was analyzed and its biochemical and physiological functions were studied based on its structure, so as to understand its specific functional mechanism and to provide an anti-inflammatory mechanism based on the three-dimensional structure of 伪 1m protein. The research and development of antioxidation and immunomodulatory drugs lay the foundation. Methods sufficient purity of human 伪 1m protein was obtained by cloning, expression and purification techniques, and then protein crystals were cultured, and the three-dimensional structure of human 伪 1m protein was analyzed by X-ray diffraction data based on its three-dimensional structure. The functional mechanism of 伪 1m protein was studied by site-directed mutation and spectroscopic analysis. Results the purity of 95% human 伪 1m protein was obtained by molecular cloning, expression and purification, and the molecular sieve (Superdex75) purification showed that 伪 1m existed as monomer and dimer in the solution. High quality single crystals suitable for X-ray diffraction were successfully obtained by screening and optimizing protein crystals, and conventional diffraction data with a resolution of 2.0 and single-wavelengthanomalous scattering data with a resolution of 3.0 were collected. The three-dimensional structure of 伪 1m protein was successfully elucidated and detected by ABTS method. The results showed that the recombinant 伪 1m protein had antioxidant activity and reductase activity.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R341
本文编号：2176624
[Abstract]:Background Lipid transport protein is a protein superfamily with 30-50 members, which has a conserved three-dimensional structure, but plays a series of different biological functions. As one of the members of lipid transport protein superfamily, 伪 1-microglobulin (伪 1m) is highly conserved and widely distributed in evolution. It has been found that 伪 1-microglobulin (伪 1m) can bind and degrade heme. It has reductase activity, antioxidant activity and immunomodulatory effect. However, its functional active sites and specific functional mechanisms are still unclear. Objective in this study, the three-dimensional structure of human 伪 1m protein was analyzed and its biochemical and physiological functions were studied based on its structure, so as to understand its specific functional mechanism and to provide an anti-inflammatory mechanism based on the three-dimensional structure of 伪 1m protein. The research and development of antioxidation and immunomodulatory drugs lay the foundation. Methods sufficient purity of human 伪 1m protein was obtained by cloning, expression and purification techniques, and then protein crystals were cultured, and the three-dimensional structure of human 伪 1m protein was analyzed by X-ray diffraction data based on its three-dimensional structure. The functional mechanism of 伪 1m protein was studied by site-directed mutation and spectroscopic analysis. Results the purity of 95% human 伪 1m protein was obtained by molecular cloning, expression and purification, and the molecular sieve (Superdex75) purification showed that 伪 1m existed as monomer and dimer in the solution. High quality single crystals suitable for X-ray diffraction were successfully obtained by screening and optimizing protein crystals, and conventional diffraction data with a resolution of 2.0 and single-wavelengthanomalous scattering data with a resolution of 3.0 were collected. The three-dimensional structure of 伪 1m protein was successfully elucidated and detected by ABTS method. The results showed that the recombinant 伪 1m protein had antioxidant activity and reductase activity.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R341
【参考文献】
相关期刊论文 前2条
1 张逸波;郑文杰;黄峙;杨芳;刘杰;陈填烽;;硒杂环化合物SPO清除DPPH和ABTS自由基的光谱学研究[J];光谱学与光谱分析;2010年07期
2 林亚静;刘志杰;龚为民;;蛋白质结构研究[J];生命科学;2007年03期
本文编号：2176624
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