烟草牻牛儿基牻牛儿基焦磷酸合成酶基因的克隆及分析

采用RACE技术,从烟叶中克隆了一个新的编码GGPPS蛋白的基因,命名为Ntggpps3,并对其进行了生物信息学分析。Ntggpps3的cDNA序列为1251 bp,包含一个长度为1092 bp的开放阅读框,编码363个氨基酸。NtGGPPS3蛋白与其他植物GGPPSs高度保守。生物信息学预测结果表明,NtGGPPS3的分子量为39.5 kD,等电点为6.28,为亲水性蛋白,N端有叶绿体信号肽。二级结构主要由螺旋和无规则卷曲组成,三级结构含有两个富含天冬氨酸的区域,并通过同源建模构建了NtGGPPS3的3D模型。进化分析表明,植物的GGPPS分为大亚基和小亚基两个分支,而且NtGGPPS3属于大亚基。

Cloning and Characterization of a New Gene Encoding Geranylgeranyl Pyrophosphate Synthase from Nicotiana tabacum

A new cDNA sequence encoding GGPPS(designated as Ntggpps3) from tobacco(Nicotiana tabacum) was cloned by RACE techniques and assayed by bioinformatics.The cDNA sequence of Ntggpps3 was of 1251 bp with an open reading frame(ORF) of 1092 bp encoding 363 amino acids.The protein NtGGPPS3 shared high similarity with other plant GGPPSs.The bioinformatics prediction showed that NtGGPPS3 might be hydrophilic with the molecular weight of 39.5 kD and isoelectric point of 6.28,and its N-terminal presented a chloroplast transit peptide.The secondary structure was mainly composed of helix and random crimps,and tertiary structure contained two classical Asp-rich motifs of GGPPS proteins which were considered as the active sites.A 3-D structure of NtGGPPS3 was constructed through homology-based modeling.Phylogenetic analysis demonstrated that plant’s GGPPSs had two branches: one was large subunit and the other was small subunit and NtGGPPS3 belongs in the former.