Zhang Xiqian, Mu Hongzhen, Ma Ting, Ding Xiangzhen, Li Zhiying, Wang Sheng
Key Laboratory of Ministry of Education for Protection and Utilization of Special Biological Resources in the Western China, Yinchuan 750021, China. E-mail:
Nan Fang Yi Ke Da Xue Xue Bao. 2015 Dec;35(12):1665-71.
To explore the mechanisms by which HFBI fusions increase recombinant fusion protein accumulation in plants.
The HFBI sequence from Trichoderma reesei was synthesized and two plant expression vectors for expression of green fluorescence protein (GFP) and GFP-HFBI were constructed. The vectors were inoculated in Nicotiana benthamiana plants through agroinfiltration, and the expression levels and mRNA accumulation levels of GFP in Nicotiana leaves were examined by Western blotting, ELISA and RT-PCR.
The HFBI fusion tag significantly enhanced the accumulation of GFP in the leaves of N. benthamiana without causing toxic effects. Endoplasmic reticulum-targeted GFP-HFBI fusion induced the formation of spherical protein particles in the plant cells.
HFBI fusions can increase the accumulation of its fusion partner in plants by forming stable protein particles, which probably shields the target protein from endogenous protease-induced degadation. HFBI fusion technology provides an alternative to improving recombinant protein expression in plants from agroinfection-compatible expression vectors.
探索热稳定真菌纤维素结合蛋白（HFBI）融合蛋白增加植物中重组融合蛋白积累的机制。
合成来自里氏木霉的HFBI序列，并构建两个用于表达绿色荧光蛋白（GFP）和GFP-HFBI的植物表达载体。通过农杆菌浸润法将载体接种到本氏烟草植株中，并用蛋白质免疫印迹法、酶联免疫吸附测定法和逆转录-聚合酶链反应检测本氏烟草叶片中GFP的表达水平和mRNA积累水平。
HFBI融合标签显著增强了GFP在本氏烟草叶片中的积累，且未产生毒性作用。内质网靶向的GFP-HFBI融合蛋白诱导植物细胞中形成球形蛋白颗粒。
HFBI融合蛋白可通过形成稳定的蛋白颗粒增加其融合伴侣在植物中的积累，这可能保护目标蛋白免受内源性蛋白酶诱导的降解。HFBI融合技术为从农杆菌感染兼容表达载体提高植物中重组蛋白表达提供了一种替代方法。
