小反刍兽疫病毒P基因的克隆及其结构与功能分析

Cloning of the Phosphoprotein Gene of Peste des Petits Ruminants and Analysis of Its Structure and Function

【目的】对小反刍兽疫病毒P基因进行克隆,并对其结构和功能的关系进行分析,为研究小反刍兽疫病毒P蛋白的功能及其在病毒增殖过程中的作用奠定基础。【方法】利用RT-PCR的方法对小反刍兽疫P基因全长进行克隆,通过生物学软件对其核苷酸序列和氨基酸序列进行了比对,利用I-TASSER进行三级结构预测,然后分析其结构和功能之间的关系。【结果】P基因和MV、CDV、RPV、DMV和PDV的P基因的相似性分别为62.9%、63.3%、64.4%、65.1%和60.4%,氨基酸的相似性分别为45.9%、46.2%、50.6%、50.3%和47.0%。三级结构表明该蛋白由3个结构域构成,中间是由多个α-螺旋构成的结构域,主要行使与L大蛋白和RNA结合的功能,两边分别是N-端和C-端结构域,C-端结构域含有3个α-螺旋,分别位于458—468aa、492—499aa和503—505aa位,主要作为N蛋白α-螺旋的结合位点。【结论】本研究成功地克隆小反刍兽疫病毒P基因,分析和预测了p蛋白的结构和功能的关系,为其进一步的生物学功能研究及其应用奠定了基础。

【Objective】 The objective of the present study was to clone the gene of PPRV phosphoprotein and analyze the structure-function relationship of phosphoprotein.【Method】 The phosphoprotein entire gene of PPRV was cloned by RT-PCR,in addition,the nucleotide and a mino acid sequences were analyzed and compared with other members of Morbillivirus through bioinformatics methods,and the protein tertiary structure was predicted using I-TASSER.【Result】 Sequence analysis showed that the homologies of nucleotide sequences between PPRV and MV,CDV,RPV,DMV and PDV were 62.9%,63.3%,64.4%,65.1%,and 60.4%,respectively,and the homologies of the deduced amino acid sequences were 45.9%,46.2%,50.6%,50.3%,and 47.0%,respectively.The tertiary structure and analysis indicated that the phosphoprotein was composed of three distinct structural domains：an N-terminal hydrophilic domain,a central domain,and a C-terminal hydrophilic domain.In addition,the central hydrophobic domain of phosphoprotein had lots of alpha helix with the potential to bind both of the large protein and RNA.The C-terminal hydrophilic domain was composed of three alpha helix,and they lay in 458-468aa,492-499aa and 503-505aa,respectively,and might play a role in their binding of alpha helix of the nucleoprotein.【Conclusion】 The phosphoprotein gene from PPRV was successfully cloned.Furthermore,the structure and function of this gene were analyzed and predicted.