Specific binding of the diphtheria tox regulatory element DtxR to the tox operator requires divalent heavy metal ions and a 9-base-pair interrupted palindromic sequence.
Specific binding of the diphtheria tox regulatory element DtxR to the tox operator requires divalent heavy metal ions and a 9-base-pair interrupted palindromic sequence.
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白喉毒素调节元件 DtxR 与毒素操纵子的特异性结合需要二价重金属离子和 9 碱基对中断的回文序列。
DOI：
10.1073/pnas.89.13.5897
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发表时间：
1992
影响因子：
11.1
通讯作者：
Murphy,JR
中科院分区：
文献类型：
--
作者：
Tao,X;Boyd,J;Murphy,JR
白喉毒素的结构基因tox由一个密切相关的棒状杆菌噬菌体家族携带;然而，tox表达的调节由白喉棒状杆菌编码的调节元件dtxR控制。dtxR的分子克隆和序列分析是最近报道的。以前的研究表明，白喉毒素操纵子的DtxR介导的调节涉及铁阻遏物复合物的形成，其特异性地结合到毒素操纵子。我们已经表达和纯化DtxR从重组大肠杆菌。免疫印迹分析表明，DtxR在两株重组大肠杆菌中均为单一的M（r）28，000蛋白条带。coli和C.白喉此外，我们证明了DtxR与白喉毒素启动子/操纵子探针的结合需要向反应混合物中加入Mn 2 +;然而，通过向反应混合物中加入螯合剂2，2 '-联吡啶、抗DtxR抗血清和过量的未标记探针可以阻断结合。从tox操纵基因中删除一个9碱基对反向重复序列会导致DtxR结合丧失。这里提出的结果表明，白喉毒素表达的DtxR的调节需要在二价重金属离子的存在下，该调节因子和毒性操作员之间的直接相互作用。
The structural gene for diphtheria toxin, tox, is carried by a family of closely related corynebacteriophages; however, the regulation of tox expression is controlled by a Corynebacterium diphtheriae-encoded regulatory element, dtxR. The molecular cloning and sequence analysis of dtxR was recently described. Previous studies have suggested that DtxR-mediated regulation of the diphtheria tox operator involves the formation of an iron-repressor complex, which specifically binds to the tox operator. We have expressed and purified DtxR from recombinant Escherichia coli. Immunoblot analysis shows DtxR to be a single M(r) 28,000 protein band in both recombinant E. coli and the C7(-) and C7hm723(-) strains of C. diphtheriae. In addition, we demonstrate that the binding of DtxR to a diphtheria tox promoter/operator probe requires the addition of Mn2+ to the reaction mixture; however, binding may be blocked by addition of the chelator 2,2'-dipyridyl, anti-DtxR antiserum, and excess unlabeled probe to the reaction mixture. Deletion of one of the 9-base-pair inverted repeat sequences from the tox operator results in a loss of DtxR binding. The results presented here demonstrate that regulation of diphtheria toxin expression by DtxR requires direct interaction between this regulatory factor and the tox operator in the presence of a divalent heavy metal ion.