Peptide dimers of the basic leucine zipper protein with non-native monomer arrangements were synthesized by using C-2 chiral templates as a synthetic dimerization module. The amino acid sequence of the peptide is derived from the DNA contact region of the basic leucine zipper protein GCN4. These peptide dimers are designed to possess different geometrical constraints from that of native GCN4 with respect to the orientation of two DNA-contacting peptides. Peptide dimers constrained at the 6th position from N-terminus recognized novel palindromic DNA sequences in which the polarity of each half-site of the parent GCN4 binding sequence is reversed. This is in contrast with dimers that are constrained at the N-terminus which failed to recognize the reversed DNA sequences. Sequence-specific recognition of these palindromic DNA sequences was confirmed by DNase I footprinting. Circular dichroism spectra-analyses revealed that dimers constrained at the 6th position bind in the helical conformation to the reversed palindromic sequences, whereas the dimers constrained at the N-terminus bind the same sequence with less helical contents. The stability of specific binding complexes was not affected by the differences in the chirality of the template. However, the stability of the half-specific complex was dramatically affected by the particular enantiomer of the template.