新型蛋白质修饰剂的合成及应用
文献类型：学位论文
| 作者 | 何明磊 |
| 学位类别 | 博士 |
| 答辩日期 | 2003 |
| 授予单位 | 中国科学院过程工程研究所 |
| 授予地点 | 中国科学院过程工程研究所 |
| 导师 | 苏志国 |
| 关键词 | 四官能团修饰剂 蛋白质 化学修饰 血红蛋白 |
| 其他题名 | Synthesis and Application of New Type of Chemical Reagents for Protein Modification |
| 学位专业 | 生物化工 |
| 中文摘要 | 化学修饰不仅是研究蛋白质的结构与功能关系的一种重要手段,也是改造蛋白质性质的一种有力工具.用化学修饰的方法对蛋白质进行改造是现代生物技术发展的一个重要方向.上世纪八十年代以来,用化学修饰血红蛋白进行血液替代品的研究和用生物相容性聚合物-聚乙二醇(PEG)共价修饰细胞因子类蛋白质的研究成了医用蛋白化学修饰研究的两大热点.该文在修饰剂的合成上做了一些工作,并用自行合成的修饰剂对几种医用蛋白质进行了初步的修饰研究.首先针对血红蛋白的化学修饰,采用天然的氨基酸L-谷氨酸和己二酸为原料合成了一种新型的四官能团修饰剂-己二酰二胺二谷氨酸,通过分析证明了合成物质的化学结构与设计结构相符.接下来该文对单甲氧基聚乙二醇的活化方式进行了探索.采用不同的中等强度的氧化剂对单甲氧基聚乙二醇(mPEG-OH)进行了氧化,生成可与蛋白质在常温或低温下反应的mPEG-CHO.在前面工作的基础上,该文对新型的PEG修饰剂-分枝型及分叉型PEG的合成进行了探索.在该文的最后部分,我们使用自行合成的几种修饰剂对几种医用蛋白质进行了初步的修饰研究.使用四官能团修饰剂和分叉型PEG对牛血红蛋白的修饰研究,获得较好的结果. |
| 英文摘要 | The development of biotechnology, especially genetic engineering provide with more and more pharmaceutical proteins. However, when being used for medical purpose, these proteins demonstrate some disadvantages, such as antigenicity, short retention time, instability, etc. Chemical modification of protein is not only an important means for research work on relationship between protein structure and its function, but also a powerful tool to improve protein properties in applications. The research of blood substitute by chemical modification of hemoglobin and protein PEGylation have become two central issues in the field of chemical modification of proteins since 1980's. The aim of this thesis is to study the process of PEG activation and to synthesize novel reagents for modification of proteins. Firstly, hexanedioyl di-glutamic acid was synthesized using L-glutamic acid and hexane di-acid. This is a new type of modification reagent with four functional groups. The chemical structure, molecular weight and the number of functional groups of end product are consistent with those of the designed molecule. This molecule was specially designed for the modification of hemoglobin. It could also be used to modify other proteins. Secondly, mPEG-aldehyde was synthesized through oxidation of mono-methoxy polyethylene glycol (mPEG-OH) using moderate oxidants. This kind of PEG was used in site-specified modification research. An important type of PEG, carboxymethylated mPEG was synthesized by catalysis oxidation method and Willimson method. Willimson method was modified by using alcohol-sodium method instead of using potassium tert-butanol method , as a result , the process was simplified and the same yield was obtained. In catalysis oxidation method, a new pressure-aerated reactor was designed to decrease the quantity of catalyst and a very high yield (>99%) of carboxymethylated mPEG was obtained. A branched PEG and a forked PEG , the two novel protein modification reagents were synthesized via methods of protection, condensation, and deprotection using lysine and glutamic acid and mPEG-mono-acid as starting materials. In this method, the most important reaction of forming branched PEG and forked PEG was carried out in one step by condensation reaction in organic phase. It makes this method remarkably different from other methods. The reaction condition was mild and had a 61% yield for the branched PEG. This method was firstly used in the synthesis of PEG and has not been found in other reports. Three patents were applied in the National Bureau of Patent of China. In last section of this thesis, several chemical reagents were used to modify some medical proteins. Four-functional modification reagent and the forked PEG were used to modify bovine hemoglobin and relatively good result was obtained. The modified hemoglobin with very narrow distribution of molecular weights were easily obtained under common reaction conditions by using these two reagents. Modified products kept good oxygen-carrying ability. rhG-CSF and rh-INF- a were also modified by linear PEG and branched PEG. The modification degree of protein was measured by Flurescamine method. Several factors that could affect modification reaction were also been probed. All these studies would lay good foundations for the chemical modification of these medical proteins. |
| 语种 | 中文 |
| 公开日期 | 2013-09-16 |
| 页码 | 135 |
| 源URL | [http://ir.ipe.ac.cn/handle/122111/1382]  |
| 专题 | 过程工程研究所_研究所（批量导入） |
| 推荐引用方式 GB/T 7714 | 何明磊. 新型蛋白质修饰剂的合成及应用[D]. 中国科学院过程工程研究所. 中国科学院过程工程研究所. 2003. |
入库方式： OAI收割
来源：过程工程研究所
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