3.3-Å resolution cryo-EM structure of human ribonucleotide reductase with substrate and allosteric regulators bound.

Ribonucleotide reductases (RNRs) convert ribonucleotides into deoxyribonucleotides, a reaction essential for DNA replication and repair. Human RNR requires two subunits for activity, the alpha subunit contains the active site, and the beta subunit houses the radical cofactor. Here, we present a 3.3-angstrom resolution structure by cryo-electron microscopy (EM) of a dATP-inhibited state of human RNR. This structure, which was determined in the presence of substrate CDP and allosteric regulators ATP and dATP, has three alpha(2) units arranged in an alpha(6) ring. At near-atomic resolution, these data provide insight into the molecular basis for CDP recognition by allosteric specificity effectors dATP/ATP. Additionally, we present lower-resolution EM structures of human a6 in the presence of both the anticancer drug clofarabine triphosphate and beta(2). Together, these structures support a model for RNR inhibition in which beta(2) is excluded from binding in a radical transfer competent position when a exists as a stable hexamer.