Despite the remarkable resistance of the nucleic acid phosphodiester backbone to degradation affording genetic stability, the P-O bond must be broken during DNA repair and RNA metabolism, among many other critical cellular processes. Nucleases are powerful enzymes that can enhance the uncatalyzed rate of phosphodiester bond cleavage by up to ∼1017-fold. Despite the most well accepted hydrolysis mechanism involving two metals (MA2+ to activate a water nucleophile and MB2+ to stabilize the leaving group), experimental evidence suggests that some nucleases can use a single metal to facilitate the chemical step, a controversial concept in the literature. The present perspective uses the case studies of four nucleases (I-PpoI, APE1, and bacterial and human EndoV) to highlight how computational approaches ranging from quantum mechanical (QM) cluster models to molecular dynamics (MD) simulations and combined quantum mechanics-molecular mechanics (QM/MM) calculations can reveal the atomic level details necessary to understand how a nuclease can use a single metal to facilitate this difficult chemistry. The representative nucleases showcase how different amino acid residues (e.g., histidine, aspartate) can fulfill the role of the first metal (MA2+) in the two-metal-mediated mechanisms. Nevertheless, differences in active site architectures afford diversity in the single-metal-mediated mechanism in terms of the metal-substrate coordination, the role of the metal, and the identities of the general acid and base. The greater understanding of the catalytic mechanisms of nucleases obtained from the body of work reviewed can be used to further explore the progression of diseases associated with nuclease (mis)activity and the development of novel nuclease applications such as disease diagnostics, gene engineering, and therapeutics.