Subunit M2 of mammalian ribonucleotide reductase. Characterization of a homogeneous protein isolated from M2-overproducing mouse cells.

Abstract

The M2 subunit of mammalian ribonucleotide reductase was purified to homogeneity from hydroxyurea-resistant, M2-overproducing mouse cells. The purification procedure involved affinity chromatography on an anti-tubulin antibody-Sepharose column and high performance gel permeation chromatography. The pure protein is a dimer of Mr = 88,000, containing stoichiometric amounts of a non-heme iron center and a tyrosyl free radical. The radical is destroyed by hydroxyurea but can readily be regenerated on incubation of the radical-free protein alone with iron-dithiothreitol in the presence of air. The ability to spontaneously regenerate the tyrosyl radical distinguishes protein M2 from the corresponding subunit of Escherichia coli ribonucleotide reductase, protein B2, but apart from that the two proteins are very similar.