The association constants for the formation of sixteen purine purine pyrimidine triple helical complexes which vary at a single common position (Z . XY where Z = A, G, C, T and XY = AT, GC, CG, TA) were determined by quantitative DNAse I footprint titration. The sequence context in which the sixteen pairings Z . XY were studied is 5’-d(AGGGGAGGGGXGGGA)-3’ within a 648 base pair DNA fragment. The association constants for the purine-rich oligonucleotides 5’-d(TGGGZGGGGTGGGGT)-3’ ranged from 2.7 x 10(5) (Z . XY = G . TA) to 8.9 x 10(7) M(-1) (Z . XY = T . AT) at 37 degrees C, pH 7,4, 50 mM tris acetate, 10 mM NaCl, and 3 mM MgCl2. For the sequences studied, T . AT is more stable than T . CG, T . GC, and T . TA by greater than or equal to 1.2 kcal mol(-1); A . AT is more stable than A . CG, A . GC, and A . TA by greater than or equal to 1.2 kcal mol(-1); and G . GC is more stable than G . AT, G . CG, and G . TA by greater than or equal to 2.1 kcal mol(-1). The triplet T . CG is greater than or equal to 1.5 kcal mol(-1) more stable than the other three bases opposite a CG base pair.