Cation-π interactions play a significant role in the stabilization of globular proteins. However, their role in collagen triple helices is less well understood and they have rarely been used in de novo designed collagen mimetic systems. In this study, we analyze the stabilizing and destabilizing effects in pairwise amino acid interactions between cationic and aromatic residues in both axial and lateral sequential relationships. Thermal unfolding experiments demonstrated that only axial pairs are stabilizing, while the lateral pairs are uniformly destabilizing. Molecular dynamics simulations show that pairs with an axial relationship can achieve a near-ideal interaction distance, but pairs in a lateral relationship do not. Arginine-π systems were found to be more stabilizing than lysine-π and histidine-π. Arginine-π interactions were then studied in more chemically diverse ABC-type heterotrimeric helices, where arginine-tyrosine pairs were found to form the best helix. This work helps elucidate the role of cation-π interactions in triple helices and illustrates their utility in designing collagen mimetic peptides.