The net energetic contribution of interhelical electrostatic attractions to coiled-coil stability has been quantitated using de novo designed synthetic coiled-coils. The synthesized model coiled-coil (EK), denoted by amino acid res-idues in positions e and g, which contains only interhelical ionic interactions without any possible (i, i + 3) and (i, i + 4) intrahelical ionic interaction, consists of two identical 35 residue polypeptide chains with a heptad repeat KgLaG-bAcLdEeKf. Three mutant coiled-coils were prepared where five Glu residues at e positions in EK were mutated to Gin residues (QK); five Lys residues at g positions were altered to Gin residues (EQ) or these mutations were effected at both positions e and g (QQ). The stabilities of the four coiled-coils were determined by measuring the ellipticities at 220 nm as a function of urea concentration at 20°C. By using a double-mutant cycle analysis it was possible to isolate the energetic contribution of interhelical ionic attractions to coiled-coil stability from the other contributions such as helical preference and hydro-phobicity. The 0.37 ± 0.01 kcal/mol of energetic contribution of one interhelical ion pair to the coiled-coil stability was obtained from three independent comparisons. This finding suggests that a large number of weak interhelical electrostatic interactions on the surface of a protein can make a substantial contribution to protein stability. In addition, the energetic contributions of a single mutation E− ↑ Q, K+↑Q, Q↑ E° and E−↑ E°were also determined (ΔΔG = 0.22, 0.26, 0.46 and 0.65 kcal/mol for the single mutations, respectively). The greater contribution of a protonated Glu residue to coiled-coil stability compared with an ionized Glu residue (0.65 kcal/mol) can outweigh the relatively smaller contribution of an interhelical ion pair (0.37 kcal/mol), which clearly explains why most coiled-coils are more stable at acidic pH compared with neutral pH even when interhelical salt bridges contribute to the coiled-coil stability at neutral pH.