RIBONUCLEASE AND ITS INHIBITOR FROM BACILLUS AMYLOLIQUENFACIENS

芽孢杆菌的胞外核糖核酸酶Barnase Amyloliquefaciens和它的细胞内抑制物Barstar是 作为研究蛋白质折叠和蛋白质折叠的模型系统 蛋白质之间的相互作用。Barnase是一种同源的 存在于原核生物和真核生物中的一组核糖核酸酶。 重组DNA技术正被应用于该项目 三个主要目标：(1)促进生产；(2)审查 基因的结构和控制序列；以及(3)剪裁 在序列中专门设计修改以进行测试 蛋白质折叠理论。 野生型Barnase基因的克隆及其致死效应 大肠杆菌或枯草芽孢杆菌可通过表达 Barstar基因位于同一质粒上。大肠杆菌质粒载体 都是针对这两种蛋白质设计的，现在都可以获得 基本上纯净量为100毫克。 Dna和氨基酸序列对于和x射线都是已知的。 棕榈酶的结构已被细化。初步研究表明 BARSTAR的结构可以通过核磁共振来解决。 至少一个不活跃的Barnase突变体形成正常 带有巴斯塔尔的情结。 一种简单的寡核苷酸定向突变方法 已经设计出了质粒DNA，并将其应用于这两个基因。

Two proteins, barnase, the extracellular ribonuclease of Bacillus amyloliquefaciens, and barstar, its intracellular inhibitor, are used as a model system for the study of protein folding and protein-protein interactions. Barnase is one of an homologous group of ribonuclease occurring in both prokaryotes and eukaryotes. Recombinant DNA techniques are being applied to the project with three major aims: (1) to facilitate production; (2) to examine the structural and control sequences of the genes; and (3) to tailor specifically designed modifications in the sequences to test theories of protein folding. The lethal effect of the cloned wild-type barnase gene in either E. coli or B. subtilis can be repressed by expression of the barstar gene placed on the same plasmid. E. coli plasmid vectors have been devised for both proteins and both can now be obtained essentially pure in 100 mg quantities. DNA and amino acid sequences are known for both and the x-ray structure of barnase has been refined. Preliminary studies suggest that the structure of barstar may be solvable by NMR. At least one Inactive barnase mutant forms a normal complex with barstar. A simple procedure for oligonucleotide-directed mutation using only plasmid DNA has been devised and applied to both genes.