以整合酶为靶点在天然药物中筛选抗HIV-1有效成分

发布时间：2018-05-13 04:40

本文选题：艾滋病 + 人类免疫缺陷病毒　；参考：《北京工业大学》2014年硕士论文

【摘要】：自1981年美国研究人员发现世界首例AIDS病例后，AIDS在全球范围内迅速蔓延,且至今尚无有效的治疗方法。它严重的威胁着人类的公共卫生安全。AIDS由人类免疫缺陷病毒引起，HIV包括两种密切相关的类型，分别为HIV-1和HIV-2，绝大多数AIDS是由HIV-1引起的。 HIV-1的复制要经过穿入、逆转录、整合、基因表达、装配、出芽和成熟等一系列步骤。其中HIV-1的整合酶在病毒的复制过程中起着十分重要的作用，它可将反转录得到的双链DNA插入宿主细胞基因当中，帮助病毒DNA与宿主DNA的融合。最后在各种病毒非结构蛋白及细胞蛋白的参与下完成病毒颗粒的复制。整合酶介导着病毒基因与宿主基因之间的融合，是寻找抗HIV药物的理想靶点。本研究的主要目的是通过一系列的研究手段从天然药物KA-60中筛选出有效地抗HIV-1整合酶单体。实验在制备高纯度HIV-1整合酶的基础上，，对天然药物KA-60进行提取，以整合酶为靶点利用SPR技术对所提取部位进行筛选，得到有效的药物提取部位。接着对所提取部位进行细胞内药效学验证。利用一种蛋白偶联柱偶联整合酶蛋白对所提取药物进行分离提纯，得到富集后的产物KA-60-6与KA-60-7。然后借助HPLC-MS对分离得到的富集产物进行分析，得到它们的准分子量并与化合物分子量数据库进行比对，筛选出药物中分子量与之相对应的物质H、A、E，并对其进行细胞内药效学检测，确定其抗HIV-1的效果。并辅以MacSynergy II软件进行有效成分联合用药水平的检测。最终，利用Autodock软件模拟有效成分与HIV-1整合酶的对接，结果显示三者均与HIV-1整合酶有一定的结合，其中单体H的结合能力最强，与药效学实验结果相符，也为之后药物结构的优化与改造奠定了基础。
[Abstract]:Since the United States researchers found the world's first case of AIDS cases in 1981, AIDS has spread rapidly around the world, and there is no effective treatment yet. It seriously threatens human public health security.AIDS caused by human immunodeficiency virus, and HIV includes two closely related types, HIV-1 and HIV-2, and the vast majority of AIDS are Caused by HIV-1.
HIV-1 replication has to pass through a series of steps such as penetration, reverse transcription, integration, gene expression, assembly, buds and maturation. In which HIV-1 integrase plays a very important role in the replication of the virus, it can insert reverse transcriptional double stranded DNA into the host cell gene and help the fusion of virus DNA and host DNA. The virus particles are replicated with the participation of non structural proteins and cell proteins. Integrase mediated fusion between the virus gene and the host gene is an ideal target for the search for anti HIV drugs.
The main purpose of this study is to screen out effective anti HIV-1 integrase monomers from natural drug KA-60 through a series of research methods. On the basis of the preparation of high purity HIV-1 integrase, the extraction of natural drug KA-60 was carried out, and the target sites were screened by SPR technology with integrated enzyme as the target, and effective drug extraction was obtained. The extracted parts were tested in cell pharmacodynamics. The extracted drugs were separated and purified with a protein coupling column coupled integrase protein. The enriched products, KA-60-6 and KA-60-7., were then analyzed with the aid of HPLC-MS, and their molecular weights and compounds were obtained. The sub quantity database was compared, and the substances corresponding to the molecular weight of the drug H, A, and E were screened out, and the intracellular pharmacodynamic test was carried out to determine the effect of its anti HIV-1. And the MacSynergy II software was used to test the level of the combined use of effective components. Finally, the Autodock software was used to simulate the docking of the effective components with the HIV-1 integrase. The results showed that both of the three were combined with the HIV-1 integrase, and the binding ability of the monomer H was the strongest, which was consistent with the results of the pharmacodynamics experiment. It also laid the foundation for the optimization and transformation of the drug structure.

【学位授予单位】：北京工业大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R96

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