We report herein the structuring of single-stranded thymine-rich DNA sequences into peptide-DNA hairpin triplex structures via designed melamine-thymine nucleobase recognition. Melamine-displaying α-peptides were synthesized with the general form (EM*)n, where M* denotes a lysine residue side chain derivatized with melamine, a bifacial hydrogen bond complement for thymine. We have found that (EM*)n peptides, which we term bifacial peptide nucleic acid (bPNA), function as a noncovalent template for thymine-rich DNA tracts. Unstructured DNA of the general form dTnCmTn are bound to (EM*)n peptides and fold into cooperatively melting 1:1 bPNA-DNA hairpin complexes with dissociation constants in the submicromolar to low nanomolar range for n = 4-10. As the length of the interface (n) is decreased, the melting temperature of the bPNA-DNA complex drops significantly, though Kd increases are less substantial, suggestive of strong enthalpy-entropy compensation. This is borne out by differential scanning calorimetry analysis, which indicates enthalpically driven bPNA-DNA base-stacking that becomes markedly less exothermic as the recognition surface n decreases in size. The recognition interface tolerates a high number of "mismatches" and indicates half-site, or monofacial, recognition between melamine and thymine may occur if only 1 complementary nucleobase is available. Association correlates directly with fractional thymine content, with optimal binding when the number of T-T sites match the number of melamine units. Interestingly, when a DNA host has more T-T sites than melamine sites on bPNA, two or three bPNAs can bind to a single DNA, resulting in ternary and quaternary complexes that have higher thermal stability than the binary (1:1) bPNA-DNA complex, suggestive of cooperative multisite binding. In contrast, when two bPNAs of different lengths bind to the same DNA host, a ternary complex is formed with two melting transitions, corresponding to independent melting of each bPNA component from the complex. These data demonstrate that melamine-displaying bPNA recognize thymine-rich DNA in predictable and multifaceted ways that allow binding affinity, structure stability, and stoichiometry to be tuned through simple bPNA length modification and matching with DNA length. Synthetic bPNA structuring elements may be useful tools for biotechnology.