Mouse ribonucleotide reductase: from genes to proteins.

Ribonucleotide reductase (EC 1.17.4.1 -2) catalyses the direct reduction of ribonucleotides to the corresponding deoxyribonucleotides. This reaction enabled the evolution from the early ‘RNA world’ to the present ‘DNA world’. Storage of genetic information as DNA instead of RNA is much more stable and this was a prerequisite for the later development of multiple, separate, species [l] . Ribonucleotide reduction is a chemically difficult reaction and requires free radical chemistry. We now know of three different classes of ribonucleotide reductases [ 11. Catalysis in all three classes is dependent on an enzyme-centred free radical, which in the class I enzymes is an iron-centre-generated tyrosyl free radical. Class I1 uses adenosyl cobalamin as a cofactor, which generates a transient radical during catalysis. Finally, class I11 uses a glycine radical which is generated in a reaction that requires S-adenosylmethionine. All plant and eukaryotic ribonucleotide reductases described so far belong to class I. Here the enzyme consists of two non-identical homodimeric subunits, proteins R1 and R2. This is very similar to the extensively studied Escherichiu coli nrd AB enzyme [2-41. The 2 x 90 kDa mouse R1 protein contains the ribonucleoside diphosphate substrate binding sites as well as two different types of nucleoside triphosphate effector binding sites. It also contains a number of redox active disulphides that participate in electron transfer from NADPH/thioredoxin/ glutaredoxin. There is only one enzyme catalysing the reduction of all four different ribonucleotides, but both the overall activity and the substrate specificity are controlled by allosteric regulation. This control ensures a balanced supply of the four deoxyribonucleotides during DNA replication and repair to prevent misincorporation and mutagenesis [3,5]. Each 45 kDa polypeptide of the mouse R2 protein contains a binuclear iron centre which, during its formation, generates a stable tyrosyl free radical that is essential for activity [6,7]. The iron centre was shown to be labile at physiological temperatures, resulting in a loss of about