Murialdo Helios, Xing Xuekun, Tzamtzis Dimitra, Haddad Abraham, Gold Marvin
Fundación Ciencia para la Vida and Millennium Institute for Fundamental and Applied Biology, Avenida Marathon 1943, Santiago, Chile.
Biochem Cell Biol. 2003 Aug;81(4):307-15. doi: 10.1139/o03-059.
Gene W is one of the 10 genes that control the morphogenesis of the bacteriophage lambda head. The morpho genesis of the phage lambda head proceeds through the synthesis of an intermediate assembly called the prohead. This is an empty shell into which the bacteriophage DNA is introduced--packaged--by the phage enzyme DNA terminase. The product of W (gpW) acts after DNA packaging, but before the addition of another phage product, gene product FII, and before the addition of tails. The role of gpW is unknown. The structure of N- and C-tagged gpW has been previously determined by nuclear magnetic resonance (NMR) spectroscopy. Here we report some of the properties of the native protein. The purification of gpW to homogeneity, overproduced by a plasmid derivative, is described. To obtain large amounts of the protein, the ribosome-binding site had to be modified, showing that inefficient translation of the message is the main mechanism limiting W gene expression. The molecular weight of the protein is in close agreement to the value predicted from the DNA sequence of the gene, which suggests that it is not post-transcriptionally modified. It behaves as a monomer in solution. Radioactively labeled gpW is incorporated into phage particles in in vitro complementation, showing that gpW is a structural protein. The stage at which gpW functions and other circumstantial evidence support the idea that six molecules of gpW polymerize on the connector before the incorporation of six molecules of gpFII and before the tail attaches.
基因W是控制λ噬菌体头部形态发生的10个基因之一。λ噬菌体头部的形态发生通过一种称为前头部的中间组装体的合成来进行。这是一个空壳，噬菌体DNA通过噬菌体酶DNA末端酶被引入其中——进行包装。W基因的产物（gpW）在DNA包装之后、另一种噬菌体产物基因产物FII添加之前以及尾部添加之前发挥作用。gpW的作用尚不清楚。N端和C端标记的gpW的结构先前已通过核磁共振（NMR）光谱确定。在此我们报告天然蛋白的一些特性。描述了通过质粒衍生物过量表达并纯化至同质的gpW。为了获得大量该蛋白，必须对核糖体结合位点进行修饰，这表明信使RNA翻译效率低下是限制W基因表达的主要机制。该蛋白的分子量与根据基因DNA序列预测的值非常一致，这表明它没有进行转录后修饰。它在溶液中表现为单体。放射性标记的gpW在体外互补实验中被整合到噬菌体颗粒中，表明gpW是一种结构蛋白。gpW发挥作用的阶段以及其他间接证据支持这样一种观点，即在六个分子的gpFII掺入之前以及尾部附着之前，六个分子的gpW在连接体上聚合。
