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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。
In order to elucidate the expression characteristics of eIF (iso) 4E, a novel transactivation factor isoform of cabbage, and its interaction with turnip mosaic virus (TuMV), eIF (iso ) 4E.a gene, the eIF (iso) 4E.c gene was cloned in ’BP8407’. The two genes are identical in length and both encode 200 amino acids. Yeast two-hybrid and bimolecular fluorescence complementation experiments showed that eIF (iso) 4E.a only interacted with TuMV-C4 lines and did not interact with TuMV-UK1 lines; eIF (iso) 4E.c only interacted with TuMV- UK1 lines interact without interacting with TuMV-C4 lines. The amino acid sequence and spatial structure of eIF (iso) 4E protein of Chinese cabbage revealed that the eIF (iso) 4E protein contained a cap-binding protein structure with 20 different amino acids between eIF (iso) 4E.a and eIF (iso) 4E.c proteins Of which 17 different amino acids were located on the capsid-binding protein structure. TuMV-C4 VPg and TuMV-UK1 VPg protein had 4 amino acid difference sites. Through the frequency analysis of the amino acid sequence of VPg protein (virus genomic connexin) in different strains of TuMV, the frequency of amino acid selection was found to be at the four amino acid difference sites.