GENETIC ANALYSIS OF THE SECRETORY APPARATUS OF E COLI

在大肠杆菌的分泌器官中的突变体，它阻止了 出口的蛋白质，已被分离。在真核生物中，它已经被证明 信号序列突变可以识别分泌物的一个成分 设备；这一承认导致在翻译中停止 出口蛋白质。直到被阻止的翻译才恢复合成 复合体与内质网(ER)膜接触。这个 大肠杆菌分泌型突变体SECC被分离为一个假回复突变体。 另一种分泌物突变，塞卡。在23摄氏度，SECC突变体阻止 运输蛋白质的合成。在这个突变体中，合成障碍 通过突变输出蛋白的信号序列可以逆转 这种蛋白质。 这项提议有几个目的。首先，出口蛋白质中的突变， 特异性地逆转了SECC中该蛋白质的合成障碍 将被分离并测序。这项工作应该定义 参与蛋白质合成的出口蛋白质的结构域。第二, 分离SECC突变体的假逆转株应识别基因 其蛋白质产物与SECC基因产物相互作用。第三， SECC基因敲除突变体的分离及表型分析 最终解开SECC蛋白的作用。这个基因敲除的突变体 将通过Tn5诱变构建。进一步的特征描述 SECC突变体也将完成。针对SecA的抗体 蛋白质将被用来确定SECC和SECA蛋白 互动。免疫背诵和温度变化实验，在 存在一种阻止蛋白质合成的药物， 应确定合成中的区块是否具有特定的位点。其效果 SECC突变对SECC蛋白的影响也将得到评估。 蛋白质分泌是细胞的一个基本过程。关于中国石油天然气集团公司 大肠杆菌的分泌机制可能会加深我们对这一点的理解 在所有有机体中都有这个过程。

Mutants in the secretory apparatus of E. coli, which block the synthesis of exported proteins, have been isolated. In eukaryotes, it has been shown that signal sequence mutations can recognize a component of the secretory apparatus; this recognition results in an arrest in the translation of exported proteins. Synthesis is not resumed until the arrested translation complex comes in contact with the endoplasmic reticulum (ER) membrane. The E. coli secretory mutant, secC, was isolated as a pseudorevertant of another secretion mutant, secA. At 23 degrees C, the secC mutant blocks the synthesis of transported proteins. In this mutant, the synthesis block of an exported protein can be reversed by mutating the signal sequence of that protein. This proposal has several aims. First, mutations in an exported protein, which specifically reverse the synthesis block of that protein in the secC mutant, will be isolated and sequenced. This work should define the domains of an exported protein that are involved in its synthesis. Second, the isolation of pseudorevertants of the secC mutant should identify genes whose protein products interact with the secC gene product. Third, the isolation and phenotypic analysis of a knock-out mutant in secC should ultimately unravel the role of the secC protein. This knock-out mutant will be constructed by using Tn5 mutagenesis. Further characterization of the secC mutant will also be done. Antibody directed against the SecA protein will be used to determine whether the SecC and SecA proteins interact. Immunoprecitation and temperature shift experiments, in the presence of a drug that prevents the initiation of protein synthesis, should determine if the block in synthesis is site-specific. The effect that the secC mutation has on the SecC proteins will also be evaluated. Protein secretion is an essential cellular process. Studies on the mechanism of secretion in E. coli may strengthen our understanding of this process in all organisms.