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目的研究人乳头瘤病毒(Human papilloma virus,HPV)58晚期蛋白1(L1)羧基端核定位信号在毕赤酵母细胞中的入核功能。方法分别构建全长HPV58L1基因和缺失羧基端核定位信号的HPV58L1基因与绿色荧光蛋白(Green fluores-cent protein,GFP)基因融合的重组表达质粒,在毕赤酵母中表达相应蛋白,用激光共聚焦显微镜观察融合蛋白在毕赤酵母细胞中的分布情况。结果融合基因表达质粒经双酶切及测序证实构建正确;表达的融合蛋白没有聚集在毕赤酵母细胞核中,而是随机地分布在毕赤酵母细胞细胞核和细胞质中。结论 HPV58 L1蛋白的羧基端核定位信号在毕赤酵母细胞中不发挥主动入核的功能。
Objective To study the nuclear import function of the carboxyl terminal signal of 58 endoprotein 1 (L1) of human papilloma virus (HPV) in Pichia pastoris cells. Methods The full length HPV58 L1 gene and HPV58 L1 gene lacking the signal of carboxyl terminal nucleus were fused with the green fluorescent protein (GFP) gene respectively. The recombinant protein was expressed in Pichia pastoris. The distribution of the fusion protein in Pichia pastoris cells was observed with a microscope. Results The fusion gene expression plasmid was confirmed by double enzyme digestion and sequencing. The expressed fusion protein did not accumulate in the Pichia pastoris nucleus, but was randomly distributed in the nucleus and cytoplasm of Pichia pastoris. Conclusion The carboxyl-terminal localization signal of HPV58 L1 protein does not play the role of active nucleation in Pichia pastoris cells.