Structural insight into operator dre-sites recognition and effector binding in the GntR/HutC transcription regulator NagR
Structural insight into operator dre-sites recognition and effector binding in the GntR/HutC transcription regulator NagR
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DOI：
10.1093/nar/gku1374
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发表时间：
2015-01-30
影响因子：
14.9
通讯作者：
Muller, Yves A.
中科院分区：
文献类型：
--
作者：
Fillenberg, Simon B.;Grau, Florian C.;Muller, Yves A.
枯草杆菌中N-乙酰葡萄糖胺（GlcNAc）的吸收和代谢由NagR（原名YvoA）控制，NagR是广泛存在的GntR/ HutC转录调节因子家族的成员。在与特定DNA操纵子位点（dre位点）结合后，NagR阻断用于GlcNAc利用的基因的转录，并且NagR与效应子的相互作用消除基因阻遏。在这里，我们报告的晶体结构的NagR在复杂的操纵基因DNA和复杂的假定效应分子葡糖胺-6-磷酸（GlcN-6P）和N-乙酰葡糖胺-6-磷酸（GlcNAc-6-P）。不同的构象状态的比较表明，效应器能够取代的NagR-DNA结合结构域（NagR-DBDs）的近70埃后结合。此外，与回文双链DNA（dsDNA）复合的分离的NagR-DBDs的高分辨率晶体结构公开了高度序列特异性操纵子dre位点识别和NagR与dsDNA的非特异性结合的决定因素。广泛的生物化学结合研究调查的亲和力的全长NagR和分离的NagR-DBD的随机DNA，dre-网站衍生的回文或自然发生的非回文dre-网站序列表明，适当的NagR功能依赖于效应诱导的微调的DNA结合亲和力的NagR，而不是在一个完全废除其DNA结合。
The uptake and metabolism of N-acetylglucosamine (GlcNAc) in Bacillus subtilis is controlled by NagR (formerly named YvoA), a member of the widely-occurring GntR/ HutC family of transcription regulators. Upon binding to specific DNA operator sites (dre-sites) NagR blocks the transcription of genes for GlcNAc utilization and interaction of NagR with effectors abrogates gene repression. Here we report crystal structures of NagR in complex with operator DNA and in complex with the putative effector molecules glucosamine-6-phosphate (GlcN-6P) and N-acetylglucosamine-6-phosphate (GlcNAc-6-P). A comparison of the distinct conformational states suggests that effectors are able to displace the NagR-DNA-binding domains (NagR-DBDs) by almost 70 angstrom upon binding. In addition, a high-resolution crystal structure of isolated NagR-DBDs in complex with palindromic double-stranded DNA (dsDNA) discloses both the determinants for highly sequence-specific operator dre-site recognition and for the unspecific binding of NagR to dsDNA. Extensive biochemical binding studies investigating the affinities of full-length NagR and isolated NagR-DBDs for either random DNA, dre-site-derived palindromic or naturally occurring non-palindromic dre-site sequences suggest that proper NagR function relies on an effector-induced fine-tuning of the DNA-binding affinities of NagR and not on a complete abrogation of its DNA binding.