Practice question

DNA synthesis proceeds via nucleophilic attack of primer 3' hydroxyl on alpha-phosphate of incoming dNTP. Two Mg2+ ions, coordinated by conserved aspartate residues in polymerase active site, orchestrate catalysis. One ion activates 3'OH lowering pKa for attack, the other stabilizes pentacovalent transition state and facilitates pyrophosphate departure. Without Mg2+, phosphodiester bond formation stalls entirely. This two-metal mechanism explains why chelators like EDTA inhibit polymerization, why Ca2+ fails to substitute effectively, and why high fidelity polymerases absolutely require magnesium for both polymerase and exonuclease activities.