Abstract
SAMHD1 is a metal-dependent deoxynucleotide triphosphohydrolase that plays a central role in regulating intracellular dNTP pools and restricting viral replication. While the structure and mechanism of human SAMHD1 (Hs SAMHD1) have been extensively characterized, its functional properties in evolutionarily distant homologs remain poorly understood. In this study, we investigate SAMHD1 proteins from the Leishmania tarentolae (trypansomatid) and Acanthamoeba polyphaga mimivirus (giant virus) to explore conservation and divergence of catalytic activity, metal dependence, and oligomerization. Our findings show that both proteins are active dNTPases but with very distinct features from those of the human enzyme. Unlike Hs SAMHD1, Lt SAMHD1 is highly selective for dCTP and activity is strongly dependent on transition metal ions, with manganese being the most potent activator.. Ap SAMHD1 demonstrates broader substrate specificity resembling that of the human enzyme, although it retains some preference of dCTP, a property that resembles the parasitic Lt SAMHD1. Ap SAMHD1 requires transition metals (Mn²⁺, Fe²⁺, and Co²⁺) with manganese being the most potent effector. Both orthologs are allosterically activated by GTP, but with a 10-50-fold higher affinity when compared to Hs SAMHD1.
Together, these findings highlight both conserved and divergent features of SAMHD1 function across evolution, revealing differences in substrate specificity, metal utilization, and tetramer stability between protist and viral orthologs and the human enzyme. This work provides new insight into the evolution of SAMHD1-mediated nucleotide regulation and expands our understanding of its biochemical diversity.