Ribonucleotide Reductase from Calf Thymus


Ribonucleotide reductase from calf thymus was separated into two nonidentical subunits (proteins M1 and M2) by chromatography on Blue Sepharose. Each subunit was inactive when assayed alone, but full activity was recovered on recombination. Protein M1 was purified to homogeneity as demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, chromatography on Ultrogel AcA 34, and sedimentation equilibrium centrifugation. In solution, protein M l behaves as a monomer (5.7 S) with a molecular weight of 84,000. Addition of dTTP leads to dimer formation (8.8 S, molecular weight 170,000), while addition of dATP induces tetramer formation (15.2 S) . In the presence of ATP, the protein exists as a mixture of dimers and tetramers. Oligomerization probably reflects conformational changes induced by the nucleotides. Binding of nucleotides to protein M1 was measured in equilibrium dialysis experiments. A maximum of 1.8 mol of dATP or 0.7 mol of dTl" was bound by 170,000 g of protein. In both cases, cooperativity of binding was observed with final dissociation constants of 3 X 10" M and 2 X M for dATP and dTTP, respectively. These data as well as competition experiments indicated the presence of two classes of effector binding sites, one specific for ATP and dATP, while the second class in addition bound dTTP and dGTP. Protein M2 was not obtained as a homogeneous preparation. It had a molecular weight of about 110,000, did not bind nucleotides, and was not influenced by nucleotides in its sedimentation behavior.

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@inproceedings{Thelander2001RibonucleotideRF, title={Ribonucleotide Reductase from Calf Thymus}, author={Lars Thelander and Staffan Eriksson and Margareta Akerman}, year={2001} }