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Ribonucleotide Reductase (RNR) is an enzyme present in all cells, its function is to reduce ribonucleotides to deoxyribonucleotides (dNDP’s). Human ribonucleotide reductase (HRR) is a validated target in cancer therapies and catalyzes the reduction of four different substrates ADP, UDP, CDP and GDP. To attain its goal, it possesses a very well-tuned allosteric regulation system. This is managed from subunit R1 of the enzyme that contains three different nucleotide binding sites; activity, effector and catalytic site. The effector site binds dNTP effectors such as ATP, dATP, dGTP and dTTP and each effector drives specificity for certain substrates. The effector site is located in the interface of two monomers, upholding dimer formation, the minimal active form of HRR. The activity site modulates overall activity of the enzyme and promotes hexamerization of the dimer. Binding of ATP to this site increases the rate of reduction of NDP’s while binding of dATP switches off the enzyme. The fundamental question regarding HRR regulation mecanism, is whether activity is a consequence of hexamerization, crosstalk between the three sites in the dimer or both. This interrogation has been posed experimentally and in the Roitberg group we have shed light on it applying molecular dynamics (MD) using the the Amber software package. Results from performing steered MD on the hexamer suggest that dATP can form more stable hexamers and includes a more buried dimer-dimer interface. Additionally, simulations performed on the dimer indicate that occupation of the activity site has effect on two specific regions of RNR that connect with the catalytic/effector site. These results suggest that activity in HRR can be dependent on both hexamerization and crosstalk between sites in the dimer. Current efforts are focused on determining which of both situations can be decisive in determining activity in the enzyme.

Pilar Buteler

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