Nuez B, Rojo F, Salas M
Centro de Biología Molecular, Consejo Superior de Investigaciones Cientificas-Universidad Autónoma de Madrid, Canto Blanco, Spain.
Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11401-5. doi: 10.1073/pnas.89.23.11401.
Transcription from the late promoter, PA3, of Bacillus subtilis phage phi 29 is activated by the viral regulatory protein p4. A kinetic analysis of the activation process has revealed that the role of protein p4 is to stabilize the binding of RNA polymerase to the promoter as a closed complex without significantly affecting further steps of the initiation process. Electrophoretic band-shift assays performed with a DNA fragment spanning only the protein p4 binding site showed that RNA polymerase could efficiently retard the complex formed by protein p4 bound to the DNA. Similarly, when a DNA fragment containing only the RNA polymerase-binding region of PA3 was used, p4 greatly stimulated the binding of RNA polymerase to the DNA. These results strongly suggest that p4 and RNA polymerase contact each other at the PA3 promoter. In the light of current knowledge of the p4 activation mechanism, we propose that direct contacts between the two proteins participate in the activation process.
枯草芽孢杆菌噬菌体phi 29晚期启动子PA3的转录由病毒调节蛋白p4激活。对激活过程的动力学分析表明，蛋白p4的作用是稳定RNA聚合酶与启动子的结合，形成封闭复合物，而不会显著影响起始过程的后续步骤。用仅跨越蛋白p4结合位点的DNA片段进行的电泳带迁移分析表明，RNA聚合酶可以有效地阻滞由与DNA结合的蛋白p4形成的复合物。同样，当使用仅包含PA3的RNA聚合酶结合区域的DNA片段时，p4极大地刺激了RNA聚合酶与DNA的结合。这些结果强烈表明，p4和RNA聚合酶在PA3启动子处相互接触。根据目前对p4激活机制的了解，我们提出这两种蛋白之间的直接接触参与了激活过程。
