白菜翻译起始因子eIF(iso)4E.a/c与芜菁花叶病毒TuMV-C4/UK1蛋白互作分析

为阐明白菜翻译起始因子异构体eIF(iso)4E的表达特性,以及该异构体与芜菁花叶病毒(TuMV)的互作关系,在白菜‘极早春’中克隆了eIF(iso)4E.a基因,在‘BP8407’中克隆了eIF(iso)4E.c基因。这两个基因长度一致,均编码200个氨基酸。酵母双杂交和双分子荧光互补试验表明:eIF(iso)4E.a仅与TuMV-C4株系互作,而不与TuMV-UK1株系互作;eIF(iso)4E.c仅与TuMV-UK1株系互作,而不与TuMV-C4株系互作。对白菜eIF(iso)4E蛋白氨基酸序列及空间结构分析发现:eIF(iso)4E蛋白包含帽结合蛋白结构,eIF(iso)4E.a和eIF(iso)4E.c蛋白间存在20个差异氨基酸,其中17个差异氨基酸位于帽结合蛋白结构上;TuMV-C4 VPg与TuMV-UK1 VPg蛋白有4个氨基酸差异位点。通过对TuMV不同株系的VPg蛋白(病毒基因组连接蛋白)氨基酸序列频率分析发现,在4个氨基酸差异位点处,氨基酸的使用频率具有选择性。

Analysis of the Protein Interaction of eIF(iso)4E.a/c with TuMV-C4/UK1 in Brassica rapa ssp.chinensis

To clarify the expression characteristics of eukaryotic initiation factors eIF(iso)4E] and the interaction relationship of eIF(iso)4E with TuMV VPg,the eIF(iso)4E.a and eIF(iso)4E.c genes were cloned from the Brassica rapa ssp.chinensis materials'Jizaochun'and'BP8407',respectively.Both of the two genes could encode 200 amino acids.The yeast two-hybridand bimolecular fluorescence complementation (BiFC) results showed that TuMV-C4 could only interact with eIF(iso)4E.a,instead of eIF(iso)4E.c;the TuMV-UK1 could only interact with eIF(iso)4E.c,instead of eIF(iso)4E.a.Analyzing the amino acid sequence and the domain tertiary structure of eIF(iso)4E protein revealed that it contained the cap-reception site.There were 20 different amino acids between eIF(iso)4E.a and eIF(iso)4E.c,17 of which were located in the cap-reception site (accounting for 85％).Through aligning the TuMV-C4/UK1 VPg amino acid sequences,four different amino acids were found;and after comparative analyzing the sequences from different TuMV strains,the frequency of the amino acids had certain selectivity in the four amino acid points.