Structural Basis for Activation of Class Ib Ribonucleotide Reductase

The class Ib ribonucleotide reductase of Escherichia coli can initiate reduction of nucleotides to deoxynucleotides with either a Mn-2(III)-tyrosyl radical (Y center dot) or a Fe-2(III)-Y center dot cofactor in the NrdF subunit. Whereas Fe-2(III)-Y center dot can self-assemble from Fe-2(II)-NrdF and O-2, activation of Mn-2(II)-NrdF requires a reduced flavoprotein, NrdI, proposed to form the oxidant for cofactor assembly by reduction of O-2. The crystal structures reported here of E. coli Mn-2(II)-NrdF and Fe-2(II)-NrdF reveal different coordination environments, suggesting distinct initial binding sites for the oxidants during cofactor activation. In the structures of Mn-2(II)-NrdF in complex with reduced and oxidized NrdI, a continuous channel connects the NrdI flavin cofactor to the NrdF Mn-2(II) active site. Crystallographic detection of a putative peroxide in this channel supports the proposed mechanism of Mn-2(III)-Y center dot cofactor assembly.