Bereswill S, Lichte F, Greiner S, Waidner B, Fassbinder F, Kist M
University of Freiburg, Institute of Medical Microbiology and Hygiene, Department of Microbiology and Hygiene, Germany.
Med Microbiol Immunol. 1999 Aug;188(1):31-40. doi: 10.1007/s004300050102.
A homologue of the ferric uptake regulator protein Fur has recently been identified within the Helicobacter pylori genome. The promoterless gene on a plasmid did partially complement a fur-negative mutant of Escherichia coli, and was strongly positive in the Fur titration assay (FURTA). The genetic and functional characterization of the complete fur homologue performed in this study revealed that the gene is conserved among H. pylori strains ( > 95% identity), and does not carry nucleotide transitions in iron-resistant mutants of H. pylori. The fur homologue on a plasmid mediated full iron-dependent ferric uptake regulator activity in the fur-deficient mutant strains H1681 and H1780 of E. coli. Immunoblot analysis revealed that Fur from H. pylori cross-reacts with antibodies raised against Fur from E. coli. The fact that inactivation of the fur gene abolished the FURTA-positive phenotype in the E. coli indicator strain H1717, indicated that this phenotype is rather caused by the encoded protein than by real Fur titration. Subcloning of the fur gene into an expression vector allowed controlled production in E. coli, and purification of a recombinant version of the H. pylori Fur protein. In summary, the results confirm the function of the H. pylori Fur homologue as iron-dependent transcriptional repressor by its ability to interact with the Fur-regulated promoters of the genes fiu and fhuF in E. coli.
最近在幽门螺杆菌基因组中发现了铁摄取调节蛋白Fur的一个同源物。质粒上无启动子的基因部分互补了大肠杆菌的fur阴性突变体，并且在Fur滴定分析（FURTA）中呈强阳性。本研究对完整的fur同源物进行的遗传和功能表征表明，该基因在幽门螺杆菌菌株中保守（>95%同一性），并且在幽门螺杆菌的耐铁突变体中不携带核苷酸转换。质粒上的fur同源物在大肠杆菌的fur缺陷突变体菌株H1681和H1780中介导了完全铁依赖性的铁摄取调节活性。免疫印迹分析表明，幽门螺杆菌的Fur与针对大肠杆菌Fur产生的抗体发生交叉反应。fur基因失活消除了大肠杆菌指示菌株H1717中的FURTA阳性表型，这一事实表明该表型相当程度上是由编码的蛋白质引起的，而非真正的Fur滴定。将fur基因亚克隆到表达载体中可在大肠杆菌中实现可控表达，并纯化重组形式的幽门螺杆菌Fur蛋白。总之，这些结果通过幽门螺杆菌Fur同源物与大肠杆菌中fiu和fhuF基因的Fur调控启动子相互作用的能力，证实了其作为铁依赖性转录阻遏物的功能。
