摘要
将λＤＮＡ的ｎｕｔＲ序列置于Ｌａｃ启动子的下游，在ｎｕｔＲ和β－半乳糖苷酶基因（ｇａｌＫ）之间插入λ噬茵体依赖ｒｈｏ的终止子ｔＲ１，使ｇａｌＫ的表达取决于Ｎ蛋白介导的转录抗终止作用。为研究ｎｕｔＲ对抗终止的影响，系统地进行了该序列中每个碱基的点突变（Ａ→Ｃ，Ｇ→Ｔ，Ｃ→Ａ及Ｔ→Ｇ）．结果表明ｂｏｘＡ中有两个碱基（位置２和５）的突变对抗终止作用是至关重要的，使抗终止效率降低了１０倍；而其他位置的改变影响甚微。ｂｏｘＡ的缺失在ｎｕｓ￣＋宿主中使抗终止数率降低了４０％，但在ｎｕｓＢ宿主中却恢复到野生型水平。ｂｏｘＢ茎环结构中，茎部顶端的碱基对及环中邻近茎部的两个碱基的突变对抗终止作用影响很大，被认为是Ｎ蛋白的识别序列．ｂｏｘＡ和ｂｏｘＢ之间的间隔序列中有两个碱基的突变几乎使抗终止作用丧失。
In this study,in order to examine the role of transcription antitermination and nut site function,we have placed the nutR region in the downstream of a lac promoter. Between nutR and gal K,theλ rho-dependent terminator,tR1,has been placed,so that the expression of gal K depends on transcription antitermination by N protein. We have systematically changed each base (A→C,G→T,C→A,or T→G) in nutR site to examine its effect on antitermination. Our results indicate that only positions 2 and 5 of the 9 base pairs of boxA were found to be critical. The point mutation of these two positions decreases N antitermination about 10- fold. Other positional changes remain functional or cause minor decrease. Deletion of boxA decreases antitermination about 40% in nus￣+ host strains. However,NusB does not exert an effect on N-dependent transcription antitermination when the boxA segment is deleted. We have also found that every base change at any position within the 5 base boxB loop is unfavorable to antitermination by the N protein of bacteriophage λ. When the T─A base pair at the top of box B stem and the stem-adjacent two bases of the loop are changed,it causes a serious prevention in antitermination. The segment may be sequence specific for N recognition. Mutation of two positons of the 8 base interval between box A and box B causes a defect in antitermination.
