The activating component of the anaerobic ribonucleotide reductase from Escherichia coli - An iron-sulfur center with only three cysteines

Class III anaerobic ribonucleotide reductase small component, named protein beta, contains a (4Fe-4S) center. its function is to mediate electron tran sfer from reduced flavodoxin to S-adenosylmethionine, required for the intr oduction of a glycyl radical in the large component, named protein or, whic h then becomes active for the reduction of ribonucleotides. By site-directe d mutagenesis we demonstrate that the three cysteines of the conserved CXXX CXXC sequence are involved in iron chelation. Such a sequence is also prese nt in the activase of the pyruvate formate-lyase and in the biotin synthase , both carrying an iron-sulfur center involved in reductive activation of S -adenosylmethionine. Even though they are able to bind iron in the (4Fe-4S) form, as shown by Mossbauer spectroscopy, the corresponding Cys to Ala mut ants are catalytically inactive. Mutation of the two other cysteines of the protein did not result in inactivation. We thus conclude that the (4Fe-4S) cluster has, in the wild type protein, only three cysteine ligands and a f ourth still unidentified ligand.