Kuhn A, Wickner W
J Biol Chem. 1985 Dec 15;260(29):15907-13.
The major coat protein (gene 8 protein) of bacteriophage M13 has been studied intensively as a model of membrane assembly, protein packing, and protein-DNA interactions. Because this protein is essential for assembly of the phage, very few mutants have been isolated. We have therefore cloned the gene 8 into a plasmid under control of the araB promoter. In the presence of arabinose, the cloned gene is expressed at a rate comparable to that in an M13-infected cell. Plasmid-derived procoat is inserted across the plasma membrane and processed to coat at a normal rate. The coat can support plaque formation by a defective M13 virus (M13am8) with an amber mutation in its procoat gene. This complementation assay was used to screen the mutagenized, cloned gene 8 for mutants which fail to make fully functional coat. Mutants were obtained which fail to synthesize procoat, which do not convert procoat to mature coat protein, or in which the coat protein is incapable of assembling into infectious virions.
噬菌体M13的主要外壳蛋白（基因8蛋白）作为膜组装、蛋白质包装和蛋白质-DNA相互作用的模型，已得到深入研究。由于这种蛋白质对于噬菌体组装至关重要，因此分离出的突变体非常少。因此，我们已将基因8克隆到受araB启动子控制的质粒中。在阿拉伯糖存在的情况下，克隆基因的表达速率与M13感染细胞中的表达速率相当。质粒衍生的前体外壳蛋白穿过质膜插入，并以正常速率加工成外壳蛋白。这种外壳蛋白可以支持带有前体外壳基因突变的缺陷型M13病毒（M13am8）形成噬菌斑。这种互补试验用于筛选诱变的克隆基因8，以寻找不能产生功能完全正常的外壳蛋白的突变体。获得了一些突变体，它们不能合成前体外壳蛋白，不能将前体外壳蛋白转化为成熟的外壳蛋白，或者其外壳蛋白不能组装成有感染性的病毒粒子。
