González-García Verónica A, Pulido-Cid Mar, Garcia-Doval Carmela, Bocanegra Rebeca, van Raaij Mark J, Martín-Benito Jaime, Cuervo Ana, Carrascosa José L
From the Structure of Macromolecules Department, Centro Nacional de Biotecnología (CSIC), Darwin 3, Cantoblanco, 28049 Madrid and.
From the Structure of Macromolecules Department, Centro Nacional de Biotecnología (CSIC), Darwin 3, Cantoblanco, 28049 Madrid and
J Biol Chem. 2015 Apr 17;290(16):10038-44. doi: 10.1074/jbc.M114.614222. Epub 2015 Feb 19.
The majority of bacteriophages protect their genetic material by packaging the nucleic acid in concentric layers to an almost crystalline concentration inside protein shells (capsid). This highly condensed genome also has to be efficiently injected into the host bacterium in a process named ejection. Most phages use a specialized complex (often a tail) to deliver the genome without disrupting cell integrity. Bacteriophage T7 belongs to the Podoviridae family and has a short, non-contractile tail formed by a tubular structure surrounded by fibers. Here we characterize the kinetics and structure of bacteriophage T7 DNA delivery process. We show that T7 recognizes lipopolysaccharides (LPS) from Escherichia coli rough strains through the fibers. Rough LPS acts as the main phage receptor and drives DNA ejection in vitro. The structural characterization of the phage tail after ejection using cryo-electron microscopy (cryo-EM) and single particle reconstruction methods revealed the major conformational changes needed for DNA delivery at low resolution. Interaction with the receptor causes fiber tilting and opening of the internal tail channel by untwisting the nozzle domain, allowing release of DNA and probably of the internal head proteins.
大多数噬菌体通过将核酸包装在同心层中,使其在蛋白质外壳(衣壳)内达到几乎结晶的浓度来保护其遗传物质。这种高度浓缩的基因组还必须在一个名为“注入”的过程中有效地注入宿主细菌。大多数噬菌体使用一种特殊的复合体(通常是尾部)来传递基因组,而不会破坏细胞完整性。噬菌体T7属于短尾噬菌体科,有一条短的、不可收缩的尾巴,由被纤维包围的管状结构形成。在这里,我们描述了噬菌体T7 DNA传递过程的动力学和结构。我们表明,T7通过纤维识别来自大肠杆菌粗糙菌株的脂多糖(LPS)。粗糙型LPS作为主要的噬菌体受体,并在体外驱动DNA注入。使用冷冻电子显微镜(cryo-EM)和单颗粒重建方法对注入后噬菌体尾部的结构表征,以低分辨率揭示了DNA传递所需的主要构象变化。与受体的相互作用导致纤维倾斜,并通过解开喷嘴结构域使内部尾部通道打开,从而允许DNA以及可能还有内部头部蛋白的释放。