SYNTHETIC AND BIOLOGICAL STUDIES OF NUCLEIC ACIDS

细菌视紫红质，紫膜的单一蛋白 嗜盐细菌，通过泵送将光能转化为化学能 质子从细胞内到细胞外。这项研究的总体目标是 研究其结构、作用机制及生物合成 细菌视紫红质的化学、物理化学、生化和遗传学研究 接近了。 紫膜中蛋白质的一级结构和取向是 现在已经知道了。人们的注意力将集中在蛋白质的三级结构上。 据推测，有七个螺旋穿过细胞膜。折叠， 单个螺旋与周围多肽环境之间的相互作用 处于不同构象状态的视网膜(例如，光适应， 细菌视紫红质的暗适应和漂白形式)将被研究。 对于结构-功能关系的研究，一个主要方法将涉及 携带细菌视紫质基因的重组DNA的应用。这个基因 将被克隆，并将在大肠杆菌中进行表达研究。如果不表达， 合成的大肠杆菌启动子将与该基因相连。或者，再加上 将研究使用各种系统的转录和翻译 用于体外生产细菌视紫红质。基因的定点突变 克隆的DNA将用于质子泵的体外重组 它现在已经在这个实验室里被解决了。在生物合成的研究中 细菌视紫红质，信使RNA的性质，合成和插入 蛋白质进入细胞膜及其调节的不同机制(通过 视网膜、光和氧气)将被研究。

Bacteriorhodopsin, the single protein of the purple membrane in extremely halophilic bacteria, transduces light energy to chemical energy by pumping protons from inside to outside of the cell. The overall aim of this research is to study the structure, mechanism of action and biosynthesis of bacteriorhodopsin by using chemical, physicochemical, biochemical and genetic approaches. The primary structure and orientation of the protein in the purple membrane is now known. Attention will be focussed on the tertiary structure of the protein. There are presumed to be seven helices which traverse the membrane. Folding, interactions between individual helices, and polypeptidic environments around the retinal in different conformational states (e.g., light-adapted, dark-adapted, and the bleached form) of bacteriorhodopsin will be investigated. For studies of structure-function relationships, a major approach will involve the use of recombinant DNA carrying the gene for bacteriorhodopsin. This gene will be cloned and its expression will be studied in E. coli. If not expressed, synthetic E. coli promoters will be linked to the gene. Alternatively, coupled transcription and translation using a variety of systems will be investigated for production of bacteriorhodopsin in vitro. Site-directed mutagenesis of the cloned DNA will be used by using in vitro reconstitution of the proton pump which now has been worked out in this laboratory. In studies of biosynthesis of bacteriorhodopsin, the nature of the messenger RNA, the synthesis and insertion of the protein into membrane and different mechanisms of its regulation (by retinal, light and oxygen) will be studied.