乙型肝炎病毒X蛋白与翻译延长因子lal（eEFlal）的相互作用及对其功能的影响

发布时间：2018-12-17 19:00

【摘要】：持续的乙型肝炎病毒(hepatitis B virus, HBV)感染可引起急慢性乙型肝炎,肝硬化,并与原发性肝细胞癌(HCC)的发生发展关系密切。全球有超过3亿的慢性感染人口,HBV致病的确切分子病理机制仍不清楚。 HBV为一种部分双链环状嗜肝DNA病毒,含4个编码病毒蛋白的开放读码框(ORF),其中一个X基因ORF编码HBV X蛋白(HBx),包含了154个氨基酸。HBx被认为是一个多功能蛋白,在病毒的感染、宿主细胞的凋亡、肝癌的发生以及参与病毒与宿主细胞相互作用的过程中都起到十分重要的作用。HBx在参与肿瘤的发生发展过程中所发挥的影响是当前HBV研究的热点。目前的研究表明,HBx的许多生物学功能需通过与其它的蛋白相互作用完成,HBx蛋白可以和许多的细胞因子相互作用,其中包括转录调控因子、细胞信号传导因子以及凋亡相关的蛋白。鉴于已经发现了种类繁多的HBx结合蛋白,我们认为可能还存在一些目前未知的能够和HBx蛋白相互作用的细胞蛋白。为更深入研究HBx的功能,本课题第一部分,我们运用免疫沉淀(Immunoprecipitation)及随后的基质辅助激光解吸电离飞行时间质谱鉴定(MALDI-TOF- MS)的方法筛选获得了五个HBx相互作用的肝细胞蛋白,其中一个为真核延长因子1a1(Eukaryotic Elongation Factor 1 alpha 1,eEF1a1)。eEF1a1又称为真核细胞翻译延长因子1a1,是一个重要的蛋白翻译因子。eEF1a-GTP催化氨酰tRNA结合到核糖体的A位点。eEF1a1不仅是翻译必须的蛋白,而且是一个重要的多功能蛋白,参与许多重要的细胞过程和疾病,包括信号传导、翻译控制、凋亡、细胞骨架组成、病毒复制及癌基因转化等。本课题的第二部分,通过GST pull-down实验和免疫共沉淀(Co-Immunoprecipitation,Co-IP)实验来进一步证实HBx蛋白与eEF1a1在细胞外和细胞内存在相互作用。 eEF1a1的功能包括参与蛋白翻译和促进F-肌动蛋白成束,本课题第三部分旨在研究HBx对eEF1a1相关功能的影响。结果提示HBx可以通过与eEF1a1结合,抑制细胞蛋白的总体翻译水平,减少F-肌动蛋白的成束,提示HBx通过与eEF1a1的相互作用,参与了HBV的致病性。
[Abstract]:Continuous hepatitis B virus (hepatitis B virus, HBV) infection can cause acute and chronic hepatitis B, liver cirrhosis, and is closely related to the occurrence and development of (HCC) in hepatocellular carcinoma. With more than 300 million chronic infections worldwide, the exact molecular pathogenetic mechanism of HBV remains unclear. HBV is a partially double-stranded DNA virus with open reading frame containing four viral proteins. One X gene, ORF encoding HBV X protein (HBx), contains 154 amino acids. HBx is considered to be a multifunctional protein. In virus infection, host cell apoptosis, The occurrence of liver cancer and the interaction between virus and host cell play a very important role. The influence of HBx in the process of tumor development is a hot topic in the research of HBV. Current studies have shown that many of the biological functions of HBx are accomplished by interacting with other proteins, and that HBx proteins interact with many cytokines, including transcriptional regulatory factors. Signal transduction factors and apoptosis-related proteins. Given that a wide variety of HBx binding proteins have been identified, we believe that there may still be some unknown cellular proteins that interact with HBx proteins. In order to study the function of HBx more deeply, the first part of this topic, Five hepatocyte proteins interacting with HBx were screened by immunoprecipitation (Immunoprecipitation) and subsequent matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF- MS). One of them is 1a1 (Eukaryotic Elongation Factor 1 alpha 1 eEF1a1). EEF1a1 is also called eukaryotic cell translation extension factor 1a1. EEF1a-GTP catalyzes the binding of aminoacyl tRNA to ribosome A site. EEF1a1 is not only a necessary protein for translation, but also an important multifunctional protein involved in many important cellular processes and diseases. These include signal transduction, translation control, apoptosis, cytoskeleton composition, viral replication and oncogene transformation. In the second part of this study, the interaction between HBx protein and eEF1a1 was further confirmed by GST pull-down and Co-Immunoprecipitation,Co-IP. The functions of eEF1a1 include protein translation and promotion of F- actin bundles. In the third part of this study, the effects of HBx on eEF1a1 related functions are studied. The results suggest that HBx can inhibit the total translation of cellular proteins and reduce the bundles of F- actin by binding to eEF1a1, suggesting that HBx is involved in the pathogenicity of HBV through the interaction with eEF1a1.
【学位授予单位】：福建医科大学
【学位级别】：博士
【学位授予年份】：2011
【分类号】：R373

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