Structural Biology Group, European Synchrotron Radiation Facility, BP 220 F-38043 Grenoble cedex 9, France.
Mol Microbiol. 2011 Mar;79(5):1260-75. doi: 10.1111/j.1365-2958.2010.07517.x. Epub 2011 Jan 5.
Fur, the ferric uptake regulator, is a transcription factor that controls iron metabolism in bacteria. Binding of ferrous iron to Fur triggers a conformational change that activates the protein for binding to specific DNA sequences named Fur boxes. In Helicobacter pylori, HpFur is involved in acid response and is important for gastric colonization in model animals. Here we present the crystal structure of a functionally active HpFur mutant (HpFur2M; C78S-C150S) bound to zinc. Although its fold is similar to that of other Fur and Fur-like proteins, the crystal structure of HpFur reveals a unique structured N-terminal extension and an unusual C-terminal helix. The structure also shows three metal binding sites: S1 the structural ZnS₄ site previously characterized biochemically in HpFur and the two zinc sites identified in other Fur proteins. Site-directed mutagenesis and spectroscopy analyses of purified wild-type HpFur and various mutants show that the two metal binding sites common to other Fur proteins can be also metallated by cobalt. DNA protection and circular dichroism experiments demonstrate that, while these two sites influence the affinity of HpFur for DNA, only one is absolutely required for DNA binding and could be responsible for the conformational changes of Fur upon metal binding while the other is a secondary site.
菌毛，铁摄取调节剂，是一种转录因子，控制细菌中的铁代谢。亚铁与 Fur 的结合触发构象变化，使蛋白质激活以结合特定的 DNA 序列，称为 Fur 盒。在幽门螺杆菌中，HpFur 参与酸反应，对于模型动物中的胃定植很重要。在这里，我们展示了一种功能活性的 HpFur 突变体（HpFur2M；C78S-C150S）与锌结合的晶体结构。尽管它的折叠类似于其他 Fur 和 Fur 样蛋白，但 HpFur 的晶体结构揭示了独特的结构化 N 端延伸和不寻常的 C 端螺旋。该结构还显示了三个金属结合位点：S1 是先前在 HpFur 中通过生物化学方法表征的结构 ZnS₄ 位点，以及在其他 Fur 蛋白中鉴定的两个锌位点。对纯化的野生型 HpFur 和各种突变体的定点突变和光谱分析表明，其他 Fur 蛋白共有的两个金属结合位点也可以被钴配位。DNA 保护和圆二色性实验表明，虽然这两个位点影响 HpFur 与 DNA 的亲和力，但只有一个位点绝对需要 DNA 结合，并且可能负责 Fur 在金属结合时的构象变化，而另一个位点是次要位点。
