O'Sullivan D J, Dowling D N, deLorenzo V, O'Gara F
Department of Microbiology, University College Cork, Ireland.
FEMS Microbiol Lett. 1994 Apr 15;117(3):327-32. doi: 10.1016/0378-1097(94)90579-7.
Introduction of a Pseudomonas iron-regulated promoter lacZ fusion (SP1) and a Pseudomonas transcriptional factor into Escherichia coli allowed expression of the promoter in this heterologous host. Evaluation of this promoter in wild-type and fur mutants of E. coli, by measuring beta-galactosidase activity, indicated that E. coli Fur can regulate the Pseudomonas promoter in response to iron starvation. Gel retardation assays suggested that purified Fur protein could interact with the SP1 promoter upstream of the transcriptional start. DNase I footprinting analysis established that Fur protected a primary 58-bp region (-50 to -106 bp). These protein/DNA interactions correlate with the observed in vivo regulation of the SP1 promoter in E. coli and indicate that Fur can functionally regulate a Pseudomonas iron-regulated promoter.
将假单胞菌铁调节启动子lacZ融合体(SP1)和假单胞菌转录因子引入大肠杆菌,使得该启动子能在这种异源宿主中表达。通过测量β-半乳糖苷酶活性,在大肠杆菌的野生型和fur突变体中对该启动子进行评估,结果表明大肠杆菌Fur可响应铁饥饿调节假单胞菌启动子。凝胶阻滞试验表明,纯化的Fur蛋白可与转录起始上游的SP1启动子相互作用。DNase I足迹分析确定Fur保护一个58bp的主要区域(-50至-106bp)。这些蛋白质/DNA相互作用与在大肠杆菌中观察到的SP1启动子的体内调节相关,表明Fur可在功能上调节假单胞菌铁调节启动子。