透明颤菌血红蛋白启动子低氧诱导重组基因在大肠杆菌中的表达研究  

Construction of an oxygen-responsive system using a VGB promoter for expression of heterologous proteins in Escherichia coli

作　　者：张晗星[1] 李十中[1] 祁庆生[2] 

机构地区：[1]清华大学核能与新能源研究院,北京100084 [2]山东大学生命科学学院,山东济南250100

出　　处：《山东大学学报（理学版）》2010年第9期122-126,共5页Journal of Shandong University(Natural Science)

基　　金：国家"十一五科技支撑项目"资助项目(2006BAD07A00)

摘　　要：设计了两段不同序列长度透明颤菌血红蛋白(VGB)的天然低氧启动子基因(Pvgb-L,Pvgb-S),分别与绿色荧光蛋白(GFP)或加强型绿色荧光蛋白(TRIGFP)基因克隆到pBluescript II SK-质粒中。根据重组菌在不同溶氧条件下荧光蛋白基因转录水平和表达量,经RT-PCR和荧光光谱分析证明,在低氧条件下,短链低氧启动子(Pvgb-S)能更有效地诱导下游基因GFP的表达,并有利于蛋白的正确折叠表达,其诱导效果受溶氧浓度影响而差异显著。在低氧培养条件下,低氧启动子Pvgb-S可有效诱导重组五碳糖代谢基因在大肠杆菌W3110(ΔpflB,ΔadhE)中的表达,提高重组菌木糖代谢,从而增加乳酸产量。We employed Pvgb, an oxygen-dependent promoter of VHb, for self-tuning regulation of green fluorescent protein （GFP） expression according to the natural transition of dissolved oxygen （DO） level during culture. A series of plasmid vectors were created using a promoter of varying size （Pvgb-L or Pvgb-S ） to drive expression of GFP. Transcription and protein accumulation were compared between each expression vector. Pvgb transcript level appears to parallel GFP protein accumulation. GFP expression in the Escherichia coli strain was very largely affected by variation of aeration environment. Strong expression of xylose genes were also observed using low aeration in recombinant E. coli W3110（ △pflB , △adhE） harboring pSK（ Pvgb , xylA , xyIB , talA, tktA ） . These vectors induced by Pvgb should be useful for overexpression of heterologous proteins and potential metabolic engineering of E. coli strains.

关 键 词：Pvgb 绿色荧光蛋白 大肠杆菌 低氧诱导 乳酸 

分 类 号：Q933[生物学—微生物学]