MOLECULAR GENETICS OF INFLUENZA VIRUS HAEMAGGLUTININ

本研究的总体目的是了解结构和 真核细胞膜蛋白功能的研究 这些分子的单个区域的分子水平。 在 去年，克隆这种基因的方法已经发展出来， 将它们重新引入哺乳动物细胞， 获得真实蛋白质在细胞表面上的表达。 在 原则上，这些技术可以应用于任何编码 从正常或恶性细胞中分离的细胞表面蛋白。 的工作 这项提议涉及一种特殊的膜蛋白--一种病毒编码的 血凝素 具体目标是：1. 将突变引入 疏水区、抗原区和糖基化位点 流感病毒血凝素。 2. 构建嵌合基因， 流感血凝素的疏水区被替换为 真核生物和细菌来源的其他膜蛋白。 3. 到 分离出大量突变和嵌合蛋白及其 野生型同源物。 这些蛋白质的各种生物学功能 将被确定并与它们的结构相关联， 必要时通过X射线晶体学进行详细分析。 4. 构建 含有信号和锚疏水区的哺乳动物载体 流感血凝素 这些载体将用于在细胞上表达 表面真核生物蛋白质，通常只在内部发现 牢房 通过这些实验和其他实验，希望找出： 这些蛋白质与细胞膜相互作用;这些分子如何 变得糖基化，以及这种糖基化可能起什么作用;如何 膜分子被引导到细胞的特定表面， 在那里，它们如何执行特定的生物功能， 抗原和免疫原。

The general aim of this research is to learn about the structure and functions of the membrane proteins of eukaryotic cells through analysis at the molecular level of the individual domains of such molecules. In the last year, methods have been developed to clone such genes, to insert them into expressing vectors, to reintroduce them into mammalian cells and to obtain expression of the authentic protein on the cell surface. In principle, these techniques can be applied to any gene coding for a cell-surface protein isolated from normal or malignant cells. The work in this proposal deals with one particular membrane protein - a virally-coded hemagglutinin. The specific aims are: 1. to introduce mutations within the hydrophobic domains, the antigenic regions and the glycosylation sites of influenza virus hemagglutinin. 2. to construct chimeric genes in which the hydrophobic regions of influenza hemagglutinin are exchanged for those of other membrane proteins of both eukaryotic and bacterial origin. 3. to isolate large quantities of mutant and chimeric proteins and their wild-type homologs. The various biological functions of these proteins will be determined and correlated with their structures, which will be analyzed in detail if necessary by X-ray crystallography. 4. to construct mammalian vectors containing the signal and anchor hydrophobic regions of influenza hemagglutinin. These vectors will be used to express on the cell surface eukaryotic proteins that are normally found only in the interior of the cell. By these and other experiments, it is hoped to find out: how such proteins interact with the membranes of the cell; how such molecules become glycosylated and what function such glycosylation may serve; how membrane molecules are directed to particular surfaces of the cell and once there, how they carry out specific biological functions and serve as antigens and immunogens.