It is considered that Hoogsteen base pairs and DNA triplex structures play important roles in cellular processes even though these structures are less than duplexes of Watson Crick base pairs. Molecular ions clearly affect the stability of DNA structures in vivo; however, the mechanisms are unknown. Here, we investigated the effects of sodium ions, choline ions, and tetramethylammonium ions on DNA triplexes using molecular dynamics simulations. We found that nonpolar interactions, which are associated with van der Waals interactions, and solvent-accessible surface area were more important than polar or electrostatic interactions in determining the affinity of a molecular cation for the DNA groove areas. The free energy gain due to a cation that fit optimally within a DNA groove was larger than the free energy loss due to the effect of dehydration. Cations with shapes complementary to that of a particular DNA groove configuration stabilized triplex formation, but cations that disturbed hydrogen bonds between DNA bases were destabilizing. These stabilizing and destabilizing mechanisms of molecular cations were also applicable to a DNA duplex composed of Watson Crick base pairs. The molecular-level view of cation interactions with DNA structures will guide the design of DNA devices, DNA-based drugs, and genetic therapies.