Department of Microbiology and Centre for Molecular Microbiology and Infection, Imperial College London, London SW7 2AZ, United Kingdom.
Proc Natl Acad Sci U S A. 2010 Feb 2;107(5):2247-52. doi: 10.1073/pnas.0907908107. Epub 2010 Jan 19.
Infection of Escherichia coli by the T7 phage leads to rapid and selective inhibition of the host RNA polymerase (RNAP)--a multi-subunit enzyme responsible for gene transcription--by a small ( approximately 7 kDa) phage-encoded protein called Gp2. Gp2 is also a potent inhibitor of E. coli RNAP in vitro. Here we describe the first atomic resolution structure of Gp2, which reveals a distinct run of surface-exposed negatively charged amino acid residues on one side of the molecule. Our comprehensive mutagenesis data reveal that two conserved arginine residues located on the opposite side of Gp2 are important for binding to and inhibition of RNAP. Based on a structural model of the Gp2-RNAP complex, we propose that inhibition of transcription by Gp2 involves prevention of RNAP-promoter DNA interactions required for stable DNA strand separation and maintenance of the "transcription bubble" near the transcription start site, an obligatory step in the formation of a transcriptionally competent promoter complex.
T7 噬菌体感染大肠杆菌会导致宿主 RNA 聚合酶（RNAP）迅速且选择性地受到抑制，而该酶是一种负责基因转录的多亚基酶，其受到一种小的（约 7kDa）噬菌体编码蛋白 Gp2 的抑制。Gp2 也是体外大肠杆菌 RNAP 的一种有效抑制剂。在这里，我们描述了 Gp2 的首个原子分辨率结构，该结构揭示了分子一侧表面暴露的带负电荷的氨基酸残基的独特连续排列。我们的全面突变数据表明，Gp2 另一侧的两个保守精氨酸残基对于与 RNAP 的结合和抑制是重要的。基于 Gp2-RNAP 复合物的结构模型，我们提出 Gp2 对转录的抑制涉及阻止 RNAP-启动子 DNA 相互作用，这些相互作用对于稳定 DNA 链分离和维持转录起始位点附近的“转录泡”是必需的，这是转录活性启动子复合物形成的必需步骤。
