【摘要】 目的:获得SARS冠状病毒小信封蛋白E(E蛋白),研究其性质,分析其可能的功能。方法:用PCR方法构建了E蛋白的质粒,在大肠杆菌中表达蛋白,用圆二色谱(CD)测定蛋白质的二级结构,用生物信息学方法预测蛋白质的二级结构和可能的功能,用分子模拟方法模建E蛋白的三维结构。结果:获得了纯度较高的E蛋白,并用CD测定了其二级结构性质,由CD谱推测了二级结构信息与生物信息学预测的基本一致。生物信息学分析表明,与其他冠状病毒相比,SARS冠状病毒的E蛋白一级结构和二级结构非常保守,跨膜区尤其保守。其功能可能与鼠科肝炎冠状病毒和哺乳动物,冠状病毒类似。结论:SARS冠状病毒E蛋白的表达成功,为研究其结构与功能以及研究SARS冠状病毒本身的性能奠定了基础;在膜环境和水溶液中,E蛋白以类似的方式折叠,但在水溶液中,其第一个β-片由于失去与膜表面的氢键作用而变成随机结构。更多还原

【Abstract】 AIM: To obtain the pure sample of SARS small envelope E protein (SARS E protein), study its properties and analyze its possible functions. METHODS: The plasmid of SARS E protein was constructed by the polymerase chain reaction (PCR), and the protein was expressed in the E coli strain. The secondary structure feature of the protein was determined by circular dichroism (CD) technique. The possible functions of this protein were annotated by bioinformatics methods, and its possible three-dimensional model was constructed by molecular modeling. RESULTS: The pure sample of SARS E protein was obtained. The secondary structure feature derived from CD determination is similar to that from the secondary structure prediction. Bioinformatics analysis indicated that the key residues of SARS E protein were much conserved compared to the E proteins of other coronaviruses. In particular, the primary amino acid sequence of SARS E protein is much more similar to that of murine hepatitis virus (MHV) and other mammal coronaviruses. The transmembrane (TM) segment of the SARS E protein is relatively more conserved in the whole protein than other regions. CONCLUSION: The success of expressing the SARS E protein is a good starting point for investigating the structure and functions of this protein and SARS coronavirus itself as well. The SARS E protein may fold in water solution in a similar way as it in membrane-water mixed environment. It is possible that β-sheet Ⅰ of the SARS E protein interacts with the membrane surface via hydrogen bonding, this β-sheet may uncoil to a random structure in water solution.更多还原