Laboratory of Transcription Biology, Center for DNA Fingerprinting and Diagnostics, Tuljaguda complex, 4-1-714 Mozamjahi Road, Nampally, Hyderabad 500 001, India.
Nucleic Acids Res. 2013 Aug;41(14):6839-56. doi: 10.1093/nar/gkt336. Epub 2013 May 22.
The conserved bacterial transcription terminator, Rho, is a potent target for bactericidal agents. Psu, a bacteriophage P4 capsid protein, is capable of inducing anti-termination to the Rho-dependent transcription termination. Knowledge of structural and mechanistic basis of this anti-termination is required to design peptide-inhibitor(s) of Rho from Psu. Using suppressor genetics, cross-linking, protein foot-printing and FRET analyses, we describe a conserved disordered structure, encompassing 139-153 amino acids of Rho, as the primary docking site for Psu. Also a neighbouring helical structure, comprising 347-354 amino acids, lining its central channel, plays a supportive role in the Rho-Psu complex formation. Based on the crystal structure of Psu, its conformation in the capsid of the P4 phage, and its interacting regions on Rho, we build an energy-minimized structural model of the Rho:Psu complex. In this model, a V-shaped dimer of Psu interacts with the two diagonally opposite subunits of a hexameric Rho, enabling Psu to form a 'lid' on the central channel of the latter. We show that this configuration of Psu makes the central channel of Rho inaccessible, and it causes a mechanical impediment to its translocase activity.
保守的细菌转录终止子 Rho 是杀菌剂的有效靶点。噬菌体 P4 衣壳蛋白 Psu 能够诱导 Rho 依赖性转录终止的反终止。为了从 Psu 设计 Rho 的肽抑制剂，需要了解这种反终止的结构和机制基础。通过抑制遗传学、交联、蛋白质足迹分析和 FRET 分析，我们描述了一个保守的无规卷曲结构，包含 Rho 的 139-153 个氨基酸，作为 Psu 的主要对接位点。另外，其中心通道沿线的一个相邻的螺旋结构，包含 347-354 个氨基酸，在 Rho-Psu 复合物形成中起辅助作用。基于 Psu 的晶体结构、其在 P4 噬菌体衣壳中的构象以及与 Rho 的相互作用区域，我们构建了 Rho:Psu 复合物的能量最小化结构模型。在该模型中，Psu 的 V 形二聚体与六聚体 Rho 的两个对角相对亚基相互作用，使 Psu 能够在后者的中心通道上形成“盖子”。我们表明，这种 Psu 的构象使 Rho 的中心通道无法进入，并对其转位酶活性造成机械障碍。
