Reduction and loss of the iron center in the reaction of the small subunit of mouse ribonucleotide reductase with hydroxyurea.
Ribonucleotide reductase is a key enzyme for DNA synthesis in living cells, and the mechanisms for its reactions with inhibitors are of interest because the inhibitors are potential antiproliferative agents. Protein R2, the small subunit of mouse ribonucleotide reductase, contains a pair of mu-oxo-bridged ferric ions and a tyrosyl free radical in each of its two polypeptide chains. Light absorption spectroscopy was used to probe the reactions of these redox centers with hydroxyurea (HU), a potent inhibitor of iron containing ribonucleotide reductases. In Escherichia coli protein R2, HU reacts with the tyrosyl radical without affecting the iron center. In contrast to the case for the E. coli protein, HU destroys the specific absorbance bands of both the iron center and the radical on a similar time scale in mouse protein R2, and this is accompanied by release of iron from the protein. Anaerobic experiments with the iron chelator bathophenanthroline present during the HU reaction indicate that the iron is released from the mouse R2 protein in the ferrous form after treatment with HU. The reduced iron center, formed by reaction of Fe2+ with mouse apoprotein R2 under anaerobic conditions, was found to be much less stable than the native Fe3+ site in the presence of suitable iron chelators. The observations are of importance for understanding the mode of action of HU on mammalian cells and for the general question of the stability of the iron center of mouse protein R2 in different redox states.
Nyholm
S
S
Department of Medical Biochemistry & Biophysics, University of Umeå, Sweden.
Thelander
L
L
Gräslund
A
A
D000818:Animals; D004926:Escherichia coli / Q000201:enzymology; D006918:Hydroxyurea / Q000378:metabolism*; D007501:Iron / Q000378:metabolism; D051379:Mice; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000378:metabolism*; D013057:Spectrum Analysis; D014443:Tyrosine / Q000378:metabolism
D011994:Recombinant Proteins; D014443:Tyrosine; D007501:Iron; D012264:Ribonucleotide Reductases; D006918:Hydroxyurea