RIBONUCLEASE AND ITS INHIBITOR FROM BACILLUS AMYLOLIQUENFACIENS

两种蛋白质，芽孢杆菌RNA酶，芽孢杆菌的胞外核糖核酸酶 解淀粉杆菌及其细胞内抑制剂芽孢杆菌RNA酶抑制剂， 用作研究蛋白质折叠的模型系统， 蛋白质相互作用 芽孢杆菌RNA酶是一种 在原核生物和真核生物中都存在的核糖核酸酶。 重组DNA技术正在应用于该项目， 三大目标：（1）促进生产;（2）审查 基因的结构和控制序列;以及（3）定制 在序列中专门设计的修饰以测试 蛋白质折叠理论 克隆的野生型芽孢杆菌RNA酶基因在两种细胞中的致死作用 E. coli或B.枯草芽孢杆菌可以通过表达 将芽孢杆菌RNA酶抑制剂基因置于同一质粒上。 E.大肠杆菌质粒载体 已经为这两种蛋白质设计了一种方法， 以100 mg的量基本上纯。 DNA和氨基酸序列都是已知的， 对芽孢杆菌RNA酶的结构进行了优化。 初步研究表明 芽孢杆菌RNA酶抑制剂的结构可以通过NMR来解析。 至少一种失活芽孢杆菌RNA酶突变体形成正常的 与Barstar合作 一种简单的寡核苷酸定向突变方法， 质粒DNA已被设计并应用于这两个基因。

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.