The substrate mechanism of ribonucleotide reductase (RNR) is studied using a hybrid DFT method (B3LYP) with large basis sets. With models for the calculations based on recently determined X-ray structures, different mechanisms of the steps leading from a ribonucleotide to a deoxyribonucleotide are investigated. It is suggested that the transformation occurs in six steps. The role of the amino acid residues present at the active site are studied in detail. Three cysteine residues are known to be necessary for the formation of a deoxyribonucleotide, but very recently a glutamic acid residue has also been shown to be essential. The calculations indicate that this glutamic acid residue, Glu441, is important both in steps 2 and 4.; In the suggested step 2, Glu441 transfers a hydrogen from the C3＇-OH group to the C2＇-OH group, and in step 4 it is proposed to participate in a cyclic transition state bridging the carbon and oxygen atoms of a keto group at C3＇. In both these steps an asparagine, Asn437, also plays an important role in reducing the barrier heights for the reactions. The rate-limiting step of the substrate reactions is suggested to be step 4, where a cysteine residue attacks the C3＇ center of the ribose ring. The disulfide bond is proposed to be formed in step 5. Dielectric. effects from the surrounding protein are very small and of almost no importance in this process.