MOLECULAR-CLONING AND DNA-SEQUENCE ANALYSIS OF A DIPHTHERIA TOX IRON-DEPENDENT REGULATORY ELEMENT (DTXR) FROM CORYNEBACTERIUM-DIPHTHERIAE
MOLECULAR-CLONING AND DNA-SEQUENCE ANALYSIS OF A DIPHTHERIA TOX IRON-DEPENDENT REGULATORY ELEMENT (DTXR) FROM CORYNEBACTERIUM-DIPHTHERIAE
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DOI：
10.1073/pnas.87.15.5968
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发表时间：
1990-08-01
影响因子：
11.1
通讯作者：
MURPHY, JR
中科院分区：
文献类型：
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作者：
BOYD, J;OZA, MN;MURPHY, JR
虽然白喉毒素的结构基因tox由一组密切相关的棒状噬菌体携带，但tox表达的调控在很大程度上是由其细菌宿主白喉棒状杆菌控制的。只有当铁成为限制生长速度的底物时，才能获得TOX基因产物的最佳产量。以前的研究表明，TOX表达的调节是通过铁结合的抑制子来实现的。为了便于从白喉隐翅虫基因组文库中克隆毒素调控元件，我们构建了毒素启动子/操纵子(ToxPO)-LacZ在大肠杆菌DH5.pha中的转录融合。我们报道了白喉毒素铁依赖调控元件dtxR的分子克隆和核酸序列，并证明了毒素-LacZ融合的β-半乳糖苷酶的表达受dtxR编码的蛋白以铁敏感的方式调节。此外，我们还表明oxPO-LacZ融合的表达不受大肠杆菌铁调节蛋白Fur的影响，并且dtxR蛋白不抑制Fur调节的外膜蛋白的表达。
Although the structural gene for diptheria toxin, tox, is carried by a family of closely related corynebacteriophages, the regulation of tox expression is controlled, to a large extent, by its bacterial host Corynebacterium diptheriae. Optimal yields of tox gene products are obtained only when iron becomes the growth-rate-limiting substrate. Previous studies suggest that regulation of tox expression is mediated through an iron-binding aporepressor. To facilitate molecular cloning of the tox regulatory element from genomic libraries of C. diphtheriae, we constructed a tox promoter/operator (toxPO)-lacZ transcriptional fusion in Escherichia coli strain DH5.alpha.. We report the molecular cloning and nucleic acid sequence of a diphtheria toxin iron-dependent regulatory element, dtxR, and demonstrate that expression of .beta.-galactosidase from the toxPO-lacZ fusions is regulated by dtxR-encoded protein in an iron-sensitive manner. In addition, we show that expression of the toxPO-lacZ fusion is not affected by the E. coli iron-regulatory protein Fur and that the dtxR protein does not inhibit expression of fur-regulated outer-membrane proteins.