Centro de Biologia Molecular e Engenharia Genética, Universidade Estadual de Campinas, Campinas, SP 13083-875, Brazil.
Antonie Van Leeuwenhoek. 2011 Mar;99(3):609-17. doi: 10.1007/s10482-010-9533-2. Epub 2010 Dec 4.
Acidithiobacillus ferrooxidans is one of the most widely used microorganisms in bioleaching operations to recover copper from low-grade copper sulfide ores. This work aimed to investigate the relative expression of genes related to the iron uptake system when A. ferrooxidans LR was maintained in contact with chalcopyrite or bornite as the sole energy source. Real-time quantitative PCR analysis revealed that the presence of bornite had no effect on the expression of seven genes related to the siderophore-mediated Fe(III) uptake system, while in the presence of chalcopyrite the expression of the genes was up-regulated. Bioinformatic analysis of the genomic region where these genes were found revealed the existence of three new putative DNA-binding sequences for the ferric iron uptake transcriptional regulator (Fur). Electrophoretic mobility shift assays demonstrated that a purified A. ferrooxidans His-tagged Fur protein was able to bind in vitro to each of these putative Fur boxes, suggesting that Fur regulated the expression of these genes. The expression of fur and two known Fur-regulated genes, mntH and dsrK, was also investigated in the presence of chalcopyrite. While the expression of fur and mntH was up-regulated, the expression of dsrK was down-regulated. The low amount of ferrous iron in the medium was probably responsible for the up-regulation of fur and the genes related to the siderophore-mediated Fe(III) uptake system when A. ferrooxidans LR was kept in the presence of chalcopyrite. A homology model of the A. ferrooxidans Fur was constructed and revealed that the putative DNA-binding surface presents conserved positively charged residues, supporting a previously suggested mode of interaction with DNA. The up-regulation of fur and the siderophore-mediated Fe(III) uptake genes, and the down-regulation of dsrK suggest that in the presence of chalcopyrite Fur acts as a transcription inducer and repressor.
氧化亚铁硫杆菌是生物浸出工艺中从低品位硫化铜矿中回收铜时最广泛使用的微生物之一。本研究旨在研究当氧化亚铁硫杆菌 LR 仅以黄铜矿或斑铜矿作为唯一能源时，铁摄取系统相关基因的相对表达情况。实时定量 PCR 分析表明，斑铜矿的存在对铁载体介导的 Fe(III)摄取系统的 7 个相关基因的表达没有影响，而在黄铜矿存在的情况下，这些基因的表达被上调。对这些基因所在基因组区域的生物信息学分析表明，存在三个新的可能的铁摄取转录调节因子 ( Fur ) DNA 结合序列。电泳迁移率变动分析表明，纯化的氧化亚铁硫杆菌 His 标记 Fur 蛋白能够体外结合到这些推定的 Fur 盒中的每一个，表明 Fur 调节这些基因的表达。在黄铜矿存在的情况下，还研究了 fur 和两个已知的 Fur 调节基因 mntH 和 dsrK 的表达。虽然 fur 和 mntH 的表达上调，但 dsrK 的表达下调。培养基中低浓度的亚铁离子可能是氧化亚铁硫杆菌 LR 在存在黄铜矿时上调 fur 和铁载体介导的 Fe(III)摄取系统相关基因表达的原因。构建了氧化亚铁硫杆菌 Fur 的同源模型，揭示了假定的 DNA 结合表面具有保守的正电荷残基，支持与 DNA 相互作用的先前提出的模式。fur 和铁载体介导的 Fe(III)摄取基因的上调，以及 dsrK 的下调表明，在黄铜矿存在的情况下， Fur 作为转录诱导剂和抑制剂发挥作用。
