Interplay of IscA and IscU in Biogenesis of Iron-Sulfur Clusters

Abstract

Increasing evidence suggests that sulfur in ubiquitous ironsulfur clusters is derived from L-cysteine via cysteine desulfurases. In Escherichia coli, the major cysteine desulfurase activity for biogenesis of iron-sulfur clusters has been attributed to IscS. The gene that encodes IscS is a member of an operon isc-SUA, which also encodes two highly conserved proteins: IscU and IscA. Previous studies suggested that both IscU and IscA may act as the iron-sulfur cluster assembly scaffold proteins. However, recent evidence indicated that IscA is an iron-binding protein that can provide iron for the iron-sulfur cluster assembly in IscU (Ding, H., Harrison, K., and Lu, J. Iron-sulfur clusters are one of the most ancient and ubiquitous redox centers in almost all living organisms (1-3). Throughout evolution, iron-sulfur clusters have become integral parts of diverse biological processes including energy conversion and the regulation of gene expression. It is now clear that biogenesis of iron-sulfur clusters is not a spontaneous process. The pioneering work by Dean's group (4) revealed that sulfur in iron-sulfur clusters is derived from L-cysteine via cysteine desulfurases, a group of pyridoxal 5-phosphate-dependent enzymes that are conserved from bacteria to humans (5-7). In Escherichia coli, there are at least three cysteine desulfurases: IscS 2 (5), SufS (8), and CSD (cysteine sulfinate desulfinase) (9, 10). Deletion of gene iscS greatly diminishes the specific activities of iron-sulfur proteins in E. coli (11,12), suggesting that IscS is the major cysteine desulfurase for biogenesis of ironsulfur clusters. Gene iscS is a member of an operon iscSUA, which also encodes two highly conserved proteins: IscU and IscA (13,14). Biochemical studies indicated that IscS catalyzes desulfurization of L-cysteine and transfers sulfane sulfur for the iron-sulfur cluster assembly in IscU via specific protein-protein interactions (15-18). Accordingly, IscU was characterized as an iron-sulfur cluster assembly scaffold protein (15-21).The function of IscA, however, still remains elusive. Previous studies suggested that IscA is an alternative iron-sulfur cluster assembly scaffold protein (22-29), because IscA, like IscU, can bind iron-sulfur clusters in the presence of ferrous iron and sulfide in vitro. On the other hand, recent studies indicated that IscA is a novel iron binding protein with an iron association constant of 2.0 -3.0 ϫ 10 19 M Ϫ1 in the presence of the thioredoxin reductase system (30) or dithiothreitol (31) and that the iron-loaded IscA can provide iron for the iron-sulfur cluster assembly in IscU (32, 33). To reconcile the two models proposed for the function of IscA, here we re-evaluated the ironsulfur cluster binding activity of IscA and IscU under physiologically relevant conditions and found that in the presence of ferrous iron, L-cysteine, and cysteine desulfurase IscS, IscU is a preferred iron-sulfur cluster assembly scaffold protein. On the other hand, when L-cysteine is not present in the incubation solution, Isc...

ErpA, an iron–sulfur (Fe–S) protein of the A-type essential for respiratory metabolism in Escherichia coli

“…Hence, we might expect a great diversity in the way bacteria make use of otherwise conserved components to carry out Fe-S biogenesis. Another challenge to the view of A-type proteins as scaffolds comes from in vitro investigations, which concluded that A-type proteins could be simple iron donors (30)(31)(32)(33)(34). Interestingly, the erpA gene is part of a chromosomal region that is particularly rich in iron uptake-related genes.…”

“…Three conserved Cys residues could act as ligands for these clusters, as indicated by mutagenesis studies and by the recently obtained 3D structure of IscA (26)(27)(28)(29). However, the proposal that IscA/SufA acts as a scaffold was challenged by in vitro studies, which suggest that A-type proteins act as iron chaperones, and by the fact that mutations in iscA or sufA did not yield major physiological defects (30)(31)(32)(33)(34)(35)(36). Noticeably, however, in A. vinelandii, iscA appeared to be required at high concentrations of oxygen (37).…”

Fe-S Cluster Assembly Pathways in Bacteria

“…Interestingly, competition experiments indicate that IscU rather than IscA preferentially forms an Fe-S cluster when both proteins are mixed simultaneously with IscS, iron, and cysteine. In contrast, IscA appears to bind iron much more tightly than IscU does if cysteine is omitted (132). A similar "iron-first" stepwise model of cluster assembly has been proposed for SufU (referred to as IscU in the cited work), based on its ability to bind iron with a high affinity (87).…”

“…Recent studies also indicated that L-cysteine could mobilize the iron center in IscA for Fe-S cluster assembly on IscU, whereas biologically related thiols such as N-acetyl-L-cysteine or reduced glutathione failed to mobilize the iron center (25). Based on these experiments, IscA was proposed to act as an iron chaperone that binds free iron for donation to IscU (132).…”

DsrR, a Novel IscA-Like Protein Lacking Iron- and Fe-S-Binding Functions, Involved in the Regulation of Sulfur Oxidation inAllochromatium vinosum

“…DsrR resembles the protein IscA, which is involved in iron-sulfur cluster maturation. IscA functions either as an iron chaperone, delivering iron ions to nascent iron-sulfur clusters built on the IscU scaffold (21,67), or as an alternative scaffold for [2Fe-2S] or [4Fe-4S] clusters (37,49). Sequence alignments of several DsrR homologues of sulfuroxidizing bacteria with IscA homologues of non-sulfur-oxidizing bacteria illustrate the sequence similarity between these proteins but also highlight the differences between them (Fig.…”