Ferric Uptake Regulator (Fur) Reversibly Binds a [2Fe-2S] Cluster to Sense Intracellular Iron Homeostasis in Escherichia Coli
iron homeostasis, iron–sulfur cluster, ferric uptake regulator (Fur), iron–sulfur protein, iron metabolism, repressor protein, Escherichia coli (E. coli), Mossbauer spectroscopy, electron paramagnetic resonance (EPR), intracellular iron homeostasis, IscA
The ferric uptake regulator (Fur) is a global transcription factor that regulates intracellular iron homeostasis in bacteria. The current hypothesis states that when the intracellular “free” iron concentration is elevated, Fur binds ferrous iron, and the iron-bound Fur represses the genes encoding for iron uptake systems and stimulates the genes encoding for iron storage proteins. However, the “iron-bound” Fur has never been isolated from any bacteria. Here we report that the Escherichia coli Fur has a bright red color when expressed in E. coli mutant cells containing an elevated intracellular free iron content because of deletion of the iron–sulfur cluster assembly proteins IscA and SufA. The acid-labile iron and sulfide content analyses in conjunction with the EPR and Mössbauer spectroscopy measurements and the site-directed mutagenesis studies show that the red Fur protein binds a [2Fe-2S] cluster via conserved cysteine residues. The occupancy of the [2Fe-2S] cluster in Fur protein is ∼31% in the E. coli iscA/sufA mutant cells and is decreased to ∼4% in WT E. coli cells. Depletion of the intracellular free iron content using the membrane-permeable iron chelator 2,2´-dipyridyl effectively removes the [2Fe-2S] cluster from Fur in E. coli cells, suggesting that Fur senses the intracellular free iron content via reversible binding of a [2Fe-2S] cluster. The binding of the [2Fe-2S] cluster in Fur appears to be highly conserved, because the Fur homolog from Hemophilus influenzae expressed in E. coli cells also reversibly binds a [2Fe-2S] cluster to sense intracellular iron homeostasis.
Journal of Biological Chemistry
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Fontenot, C.R., Tasnim, H., Valdes, K.A., Popescu, C.V., Ding, H. Ferric Uptake Regulator (Fur) Reversibly Binds a [2Fe-2S] Cluster to Sense Intracellular Iron Homeostasis in Escherichia Coli. Journal of Biological Chemistry. 2020, 295 (46), 15454-15463. 0DOI: 10.1074/jbc.RA120.014814