Walter Monika, Fiedler Christian, Grassl Renate, Biebl Manfred, Rachel Reinhard, Hermo-Parrado X Lois, Llamas-Saiz Antonio L, Seckler Robert, Miller Stefan, van Raaij Mark J
Physical Biochemistry, University of Potsdam, Karl-Liebknecht-Str. 24-25, Haus 25, D-14476 Potsdam-Golm, Germany.
J Virol. 2008 Mar;82(5):2265-73. doi: 10.1128/JVI.01641-07. Epub 2007 Dec 12.
A new Salmonella enterica phage, Det7, was isolated from sewage and shown by electron microscopy to belong to the Myoviridae morphogroup of bacteriophages. Det7 contains a 75-kDa protein with 50% overall sequence identity to the tail spike endorhamnosidase of podovirus P22. Adsorption of myoviruses to their bacterial hosts is normally mediated by long and short tail fibers attached to a contractile tail, whereas podoviruses do not contain fibers but attach to host cells through stubby tail spikes attached to a very short, noncontractile tail. The amino-terminal 150 residues of the Det7 protein lack homology to the P22 tail spike and are probably responsible for binding to the base plate of the myoviral tail. Det7 tail spike lacking this putative particle-binding domain was purified from Escherichia coli, and well-diffracting crystals of the protein were obtained. The structure, determined by molecular replacement and refined at a 1.6-A resolution, is very similar to that of bacteriophage P22 tail spike. Fluorescence titrations with an octasaccharide suggest Det7 tail spike to bind its receptor lipopolysaccharide somewhat less tightly than the P22 tail spike. The Det7 tail spike is even more resistant to thermal unfolding than the already exceptionally stable homologue from P22. Folding and assembly of both trimeric proteins are equally temperature sensitive and equally slow. Despite the close structural, biochemical, and sequence similarities between both proteins, the Det7 tail spike lacks both carboxy-terminal cysteines previously proposed to form a transient disulfide during P22 tail spike assembly. Our data suggest receptor-binding module exchange between podoviruses and myoviruses in the course of bacteriophage evolution.
从污水中分离出一种新的肠炎沙门氏菌噬菌体Det7，通过电子显微镜观察表明它属于噬菌体的肌尾噬菌体科形态群。Det7含有一种75 kDa的蛋白质，其整体序列与短尾噬菌体P22的尾刺内鼠李糖苷酶有50%的同源性。肌尾噬菌体与细菌宿主的吸附通常由附着在收缩尾上的长短尾丝介导，而短尾噬菌体不含尾丝，而是通过附着在非常短的非收缩尾上的粗短尾刺与宿主细胞结合。Det7蛋白的氨基末端150个残基与P22尾刺缺乏同源性，可能负责与肌尾噬菌体尾部的基板结合。从大肠杆菌中纯化出缺乏这种假定颗粒结合结构域的Det7尾刺，并获得了该蛋白的良好衍射晶体。通过分子置换确定并在1.6 Å分辨率下精修的结构与噬菌体P22尾刺的结构非常相似。用八糖进行的荧光滴定表明，Det7尾刺与其受体脂多糖的结合比P22尾刺稍松。Det7尾刺甚至比已经异常稳定的P22同源物更耐热解折叠。两种三聚体蛋白的折叠和组装对温度同样敏感且同样缓慢。尽管两种蛋白在结构、生化和序列上有密切的相似性，但Det7尾刺缺乏先前提出在P22尾刺组装过程中形成瞬时二硫键的羧基末端半胱氨酸。我们的数据表明，在噬菌体进化过程中，短尾噬菌体和肌尾噬菌体之间存在受体结合模块的交换。
