In class II transcription activation, the transcription factor normally binds to the promoter near the -35 position and contacts the domain 4 of σ factors (σ4 ) to activate transcription. However, σ4 of σ70 appears to be poorly folded on its own. Here, by fusing σ4 with the RNA polymerase β-flap-tip-helix, we constructed two σ4 chimera proteins, one from σ70 σ 4 70 c and another from σS σ 4 S c of Klebsiella pneumoniae. The two chimera proteins well folded into a monomeric form with strong binding affinities for -35 element DNA. Determining the crystal structure of σ 4 S c in complex with -35 element DNA revealed that σ 4 S c adopts a similar structure as σ4 in the Escherichia coli RNA polymerase σS holoenzyme and recognizes -35 element DNA specifically by several conserved residues from the helix-turn-helix motif. By using nuclear magnetic resonance (NMR), σ 4 70 c was demonstrated to recognize -35 element DNA similar to σ 4 S c . Carr-Purcell-Meiboom-Gill relaxation dispersion analyses showed that the N-terminal helix and the β-flap-tip-helix of σ 4 70 c have a concurrent transient α-helical structure and DNA binding reduced the slow dynamics on σ 4 70 c . Finally, only σ 4 70 c was shown to interact with the response regulator PmrA and its promoter DNA. The chimera proteins are capable of -35 element DNA recognition and can be used for study with transcription factors or other factors that interact with domain 4 of σ factors.

中文翻译：

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σ4嵌合蛋白识别-35元素的结构基础及其与PmrA反应调节剂的相互作用。

在II类转录激活中，转录因子通常在-35位附近与启动子结合，并与σ因子（σ4）的结构域4接触以激活转录。但是，σ70的σ4似乎折叠得很差。在这里，通过将σ4与RNA聚合酶β-flap-tip-helix融合，我们构建了两个σ4嵌合蛋白，一个来自肺炎克雷伯菌的σ70σ4 70 c，另一个来自σSσ4 S c。两种嵌合蛋白很好地折叠成具有对-35元素DNA的强结合亲和力的单体形式。确定与-35元素DNA结合的σ4 S c的晶体结构，发现σ4 S c采用与大肠杆菌RNA聚合酶σS全酶中的σ4相似的结构，并通过螺旋中的几个保守残基特异性识别-35元素DNA。螺旋形状。通过使用核磁共振（NMR），证明σ4 70 c可以识别与σ4 S c相似的-35元素DNA。Carr-Purcell-Meiboom-Gill弛豫分散分析表明，σ4 70 c的N末端螺旋和β-flap-tip-helix同时具有瞬时α螺旋结构，DNA结合降低了σ4 70的缓慢动力学C 。最后，仅σ4 70 c被证明与反应调节因子PmrA及其启动子DNA相互作用。嵌合蛋白能够识别-35个元素的DNA，可用于与转录因子或其他与σ因子4结构域相互作用的因子的研究。Carr-Purcell-Meiboom-Gill弛豫分散分析表明，σ4 70 c的N末端螺旋和β-flap-tip-helix同时具有瞬时α螺旋结构，DNA结合降低了σ4 70的缓慢动力学C 。最后，仅σ4 70 c被证明与反应调节因子PmrA及其启动子DNA相互作用。嵌合蛋白能够识别-35个元素的DNA，可用于与转录因子或其他与σ因子4结构域相互作用的因子的研究。Carr-Purcell-Meiboom-Gill弛豫分散分析表明，σ4 70 c的N末端螺旋和β-flap-tip-helix同时具有瞬时α螺旋结构，DNA结合降低了σ4 70的缓慢动力学C 。最后，仅σ4 70 c被证明与反应调节因子PmrA及其启动子DNA相互作用。嵌合蛋白能够识别-35个元素的DNA，可用于与转录因子或其他与σ因子4结构域相互作用的因子的研究。