Reaction Mechanism and Metal Selectivity of Human SAMHD1 Elucidated by QM/MM Calculations

2'-deoxynucleoside-5'-triphosphate triphosphohydrolases (dNTPases) constitute a crucial enzyme family that plays a pivotal role in antiviral innate immunity. Among these enzymes, human SAMHD1 has emerged as a novel dNTPase, demonstrating special catalytic structure and property. This metalloenzyme regulates cellular dNTP concentration through its ability to hydrolyze all four canonical dNTPs into their corresponding 2'-deoxynucleosides and inorganic triphosphates, a reaction requiring coordinated iron and magnesium ions for enzymatic activity. In the present work, molecular dynamics (MD) simulations and quantum mechanics/molecular mechanics (QM/MM) calculations are employed to investigate the mechanistic details of dATP hydrolysis mediated by two metal ions. Starting from the resolved crystal structure, Model1, containing a Fe2+ in the active site, was constructed. Our calculations demonstrate that SAMHD1 employs a bridging hydroxide anion OH− to attack Pα site of dNTP, triggering the cleavage of Pα–O5' bond via a trigonal-bipyramidal transition state. Simultaneously, His215 donates a proton to O5' of the leaving group, leading to the formation of 2'-deoxyadenosine and triphosphate. It is further demonstrated that the native Fe2+-Mg2+ bi-metallic centre help catalyse this hydrolysis reaction with a barrier of 13.4 kcal/mol, while the substitution from Fe2+ to Fe3+ abolishes the catalytic activity of SAMHD1. The comparisons between different QM/MM models highlight the high affinity of SAMHD1 for Fe2+ relative to Mn2+ and Mg2+ at one of bi-metallic sites. In addition, the metal ion swapping between Fe2+ and Mg2+ from their crystallographic positions is shown to elevate the energy of the reactant state, underscoring the critical influence of metal coordination geometry on catalytic activity. These computational insights not only expand the understanding of how SAMHD1 wisely modulates catalytic reactivity and metal selectivity by binding suitable metal ions, but also provide valuable foundation for guiding the design of novel drugs for antiviral therapies.