The IscA from Acidithiobacillus ferrooxidans is an iron–sulfur protein which assemble the [Fe4S4] cluster with intracellular iron and sulfur

IscA was proposed to be involved in the iron–sulfur cluster assembly in Acidithiobacillus ferrooxidans encoded by the iscSUA operon, but the role of IscA in the iron–sulfur cluster assembly still remains controversial. In this study, the IscA from A. ferrooxidans ATCC 23270 was successfully expressed in Escherichia coli, and purified by affinity chromatography to homogeneity. To our surprise, the purified IscA was observed to be an iron–sulfur protein according to MALDI-TOF-MS and spectra results, which was capable of recruiting intracellular iron and sulfur and hosted a stable [Fe4S4] cluster. Site-directed mutagenesis for the protein revealed that Cys35, Cys99 and Cys101 were in ligating with the [Fe4S4] cluster. The [Fe4S4] cluster could be assembled in apoIscA with Fe2+ and sulfide in vitro. The IscA from A. ferrooxidans may function as a scaffold protein for the pre-assembly of Fe–S cluster and then transfer it to target proteins in A. ferrooxidans.