Morphogenesis of Bacteriophage T4

在过去三年的NSF支持中， 了解病毒组装中形状和大小的调节 使用噬菌体T4。 关于头长调节的研究 显示病毒内的DNA被选择用于包装， 可以通过参考以下内容来解释的特定长度 二十面体表面晶格理论 T4中的DNA包装是 因此，这取决于衣壳所使用的设计原理， 设置其长度参数。 脉冲场电泳的应用 DNA的分离提供了最高的分辨率， T4 DNA分子。 这里提出的新工作将集中在如何测量系统 决定着DNA包装的运作 噬菌体T4的头部是 细长的，不是大多数病毒的二十面体。 突变 内部蛋白质核心或支架蛋白质都导致头部 回复到二十面体的形状。 所有导致 细长的头部（称为巨人）位于基因编码， 病毒外壳蛋白（gp 23）。 我们将检验gp23 单独控制头部伸长使用克隆的基因， 表达载体中的支架和外壳蛋白，定点 缺失和诱变以及分子结构测定。 将纯化的gp23置于结晶条件下， X射线结构测定。 T4股骨头的四种型号中 已经提出的长度确定，据信两个 可以消除（简单的模板和动力学模型）。 这将进一步证实初步数据， 长度由两种蛋白质的组合控制 游标机构和稳定的累积应变在 二十面体顶点 总之，拟议中的实验将有助于 在理解形状和大小的机制方面 控制，和DNA包装，在亚细胞水平。

During the past three years of NSF support, progress was made in understanding the regulation of shape and size in virus assembly using bacteriophage T4. Studies on head length regulation have shown that the DNA inside the virus is selected for packaging at specific lengths that can be interpreted by reference to icosahedral surface lattice theory. DNA packaging in T4 is therefore dependent on the design principles used by the capsid in setting its length parameters. Use of pulsed-field electrophoresis of DNA has provided the highest resolution yet for the separation of T4 DNA molecules. The new work proposed here will focus on how the measuring system that determines DNA packaging operates. The head of phage T4 is elongated, not icosahedral as for most viruses. Mutations in the internal protein core or scaffold proteins all cause the head to revert to its icosahedral form. All mutations that caused elongated heads (called giants) are located in the gene coding for the virus coat protein (gp 23). We will test the idea that gp 23 alone controls head elongation using cloned genes for both the scaffold and the coat protein in expression vectors, site-directed deletions and mutagenesis, and molecular structure determination. Purified gp23 will be subjected to crystallization conditions for x-ray structure determination. Of the four models for T4 head length determination that have been proposed, it is believed two can be eliminated (the simple template and the kinetic models). This will now extend and confirm preliminary data that suggest that length is controlled by a combination of a two-component protein vernier mechanism and stabilization of accumulated strain at the icosahedral vertices. Together, the proposed experiments will help substantially in understanding of the mechanisms of shape and size control, and DNA packaging, at the subcellular level.