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首页> 外文期刊>Infection and immunity >Characterization of an iron-dependent regulatory protein (IdeR) of Mycobacterium tuberculosis as a functional homolog of the diphtheria toxin repressor (DtxR) from Corynebacterium diphtheriae.
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Characterization of an iron-dependent regulatory protein (IdeR) of Mycobacterium tuberculosis as a functional homolog of the diphtheria toxin repressor (DtxR) from Corynebacterium diphtheriae.

机译：结核分枝杆菌铁依赖性调节蛋白（IdeR）作为白喉棒状杆菌白喉毒素阻遏物（DtxR）的功能同源物的特征。

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The DtxR protein from Corynebacterium diphtheriae is an iron-dependent repressor that regulates transcription from the tox, IRP1, and IRP2 promoters. A gene from virulent Mycobacterium tuberculosis H37Rv was recently shown to encode a protein, here designated iron-dependent regulator (IdeR), that is almost 60% homologous to DtxR from C. diphtheriae. A 750-bp PCR-derived DNA fragment carrying the M. tuberculosis ideR allele was subcloned to both high- and low-copy-number vectors. In Escherichia coli, transcription from the C. diphtheriae tox, IRP1, and IRP2 promoters was strongly repressed by ideR under high-iron conditions, and ideR restored normal iron-dependent expression of the corynebacterial siderophore in the C. diphtheriae dtxR mutant C7(beta)hm723. The M. tuberculosis IdeR protein was overexpressed in E. coli and purified to near homogeneity by nickel affinity chromatography. Gel mobility shift experiments revealed that IdeR bound to a DNA fragment that carried the C. diphtheriae tox promoter/operator sequence. DNAse I footprint analysis demonstrated that IdeR, in the presence of Cd2+, Co2+, Fe2+, Mn2+, Ni2+, or Zn2+, protected an approximately 30-bp region on DNA fragments carrying the tox, IRP1, or IRP2 promoter/operator sequences. IdeR reacted very weakly in Western blots (immunoblots) with antiserum against the C. diphtheriae DtxR protein, suggesting that the immunodominant epitopes of DtxR may be located in its poorly conserved carboxyl-terminal domain.
机译：白喉棒状杆菌中的DtxR蛋白是铁依赖性阻遏物，可调节来自毒素，IRP1和IRP2启动子的转录。最近显示，来自强毒结核分枝杆菌H37Rv的基因编码一种蛋白质，此处称为铁依赖性调节剂（IdeR），与白喉衣原体的DtxR几乎有60％的同源性。携带结核分枝杆菌ideR等位基因的750 bp PCR衍生的DNA片段被亚克隆到高拷贝数和低拷贝数的载体上。在大肠埃希氏菌中，ideR在高铁条件下强烈抑制白喉衣原体毒素，IRP1和IRP2启动子的转录，并且ideR恢复了白喉衣原体dtxR突变体C7（beta）中棒状铁载体的正常铁依赖性表达。 hm723。结核分枝杆菌IdeR蛋白在大肠杆菌中过表达，并通过镍亲和层析纯化至接近均一。凝胶迁移率变化实验表明，IdeR与携带白喉衣原体毒素启动子/操纵子序列的DNA片段结合。 DNAse I足迹分析表明，在Cd2 +，Co2 +，Fe2 +，Mn2 +，Ni2 +或Zn2 +存在的情况下，IdeR保护带有Tox，IRP1或IRP2启动子/操纵子序列的DNA片段上大约30 bp的区域。 IdeR在Western印迹（免疫印迹）中与抗白喉衣原体DtxR蛋白的抗血清反应非常弱，表明DtxR的免疫优势表位可能位于其保守性差的羧基末端结构域中。

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