Gansz A, Kruse U, Rüger W
Arbeitsgruppe Molekulare Genetik, Lehrstuhl Biologie der Mikroorganismen, Ruhr-Universität Bochum, FRG.
Mol Gen Genet. 1991 Mar;225(3):427-34. doi: 10.1007/BF00261683.
Gene product 33 of phage T4 is known to be essential in late transcription. Upstream from gene 33 and overlapping its 5' terminal sequence by 20 bp, we identified an open reading frame coding for a binding protein for double-stranded DNA (DsbA). Gene product DsbA is composed of 89 amino acid residues with a Mr of 10376 kDa. We purified this protein to homogeneity from over-expressing cells. Gel retardation assays reveal that it binds to DNA and footprint analyses disclose that it interacts preferentially with T4 late promoter regions. At the sites of binding the protein introduces nicks in double-stranded DNA. In vitro transcription assays performed with T4 late modified RNA polymerase on restriction fragments harbouring a T4 late promoter region prove that gene product DsbA enhances transcription from these promoter regions in the presence of gene product 33. Gene dsbA is distinct from gene das which maps close to this genomic region.
已知噬菌体T4的基因产物33在晚期转录中至关重要。在基因33上游且与它的5'末端序列重叠20个碱基对处,我们鉴定出一个编码双链DNA结合蛋白(DsbA)的开放阅读框。基因产物DsbA由89个氨基酸残基组成,分子量为10376 kDa。我们从过表达细胞中将该蛋白纯化至同质。凝胶阻滞分析表明它能与DNA结合,足迹分析揭示它优先与T4晚期启动子区域相互作用。在结合位点,该蛋白会在双链DNA上引入切口。用T4晚期修饰的RNA聚合酶对含有T4晚期启动子区域的限制性片段进行的体外转录分析证明,在基因产物33存在的情况下,基因产物DsbA能增强这些启动子区域的转录。基因dsbA与定位在该基因组区域附近的基因das不同。