Iron-sulfur cluster biosynthesis: characterization of Escherichia coli CYaY as an iron donor for the assembly of [2Fe-2S] clusters in the scaffold IscU.
The biogenesis of iron-sulfur [Fe-S] clusters requires the coordinated delivery of both iron and sulfide. Sulfide is provided by cysteine desulfurases that use L-cysteine as sulfur source. So far, the physiological iron donor has not been clearly identified. CyaY, the bacterial ortholog of frataxin, an iron binding protein thought to be involved in iron-sulfur cluster formation in eukaryotes, is a good candidate because it was shown to bind iron. Nevertheless, no functional in vitro studies showing an involvement of CyaY in [Fe-S] cluster biosynthesis have been reported so far. In this paper we demonstrate for the first time a specific interaction between CyaY and IscS, a cysteine desulfurase participating in iron-sulfur cluster assembly. Analysis of the iron-loaded CyaY protein demonstrated a strong binding of Fe(3+) and a weak binding of Fe(2+) by CyaY. Biochemical analysis showed that the CyaY-Fe(3+) protein corresponds to a mixture of monomer, intermediate forms (dimer-pentamers), and oligomers with the intermediate one corresponding to the only stable and soluble iron-containing form of CyaY. Using spectroscopic methods, this form was further demonstrated to be functional in vitro as an iron donor during [Fe-S] cluster assembly on the scaffold protein IscU in the presence of IscS and cysteine. All of these results point toward a link between CyaY and [Fe-S] cluster biosynthesis, and a possible mechanism for the process is discussed.
Layer
Gunhild
G
Laboratoire de Chimie et Biochimie des Centres Rédox Biologiques, DRDC/CB, Commissariat à l'Energie Atomique/CNRS/Université Joseph Fourier, CEA-Grenoble, UMR 5047, 17 Avenue des Martyrs, 38054 Grenoble Cedex 09, France.
Ollagnier-de Choudens
Sandrine
S
Sanakis
Yiannis
Y
Fontecave
Marc
M
D001426:Bacterial Proteins / Q000737:chemistry*; D013437:Carbon-Sulfur Lyases / Q000737:chemistry; D002474:Cell-Free System; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D007501:Iron / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D011485:Protein Binding; D013053:Spectrophotometry; D013455:Sulfur / Q000737:chemistry; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; C120906:CyaY protein, E coli; D029968:Escherichia coli Proteins; D007506:Iron-Sulfur Proteins; C411665:IscU protein, E coli; D013455:Sulfur; D007501:Iron; D013437:Carbon-Sulfur Lyases; C080546:cysteine desulfurase