Spectroscopic and crystallographic data are presented for a series of tetrapeptides and analogous depsipeptides that can form a minimal beta-hairpin (two intramolecular hydrogen bonds). These model compounds have been used to test the hypothesis that "mirror image" beta-turns promote beta-hairpin formation. This hypothesis was inspired by a statistical survey of beta-hairpins in globular proteins (Sibanda, B. L.; Thornton, J. M. Nature 1985, 316, 170), which showed that mirror image beta-turns (type I’ and type II’), although rare in general, are very commonly associated with beta-hairpins containing a two-residue loop between the strand segments. Each of our four-residue molecules contains proline at the second position, to promote a central beta-turn. The beta-turn is induced to be either "common" or "mirror-image", relative to the outer residues, by choice of residue configuration (L vs D). In methylene chloride, end-capped tetrapeptide Ac-L-Val-D-Pro-D-Ala-L-Leu-NMe(2) folds largely into the beta-hairpin conformation, while the diastereomer Ac-L-Val-L-Pro-L-Ala-L-Leu-NMe(2) displays little or no beta-hairpin folding. For each diastereomer, the hydrogen-bonded driving force for beta-hairpin folding is identical, and the dramatic difference in folding behavior therefore reflects a variation in the intrinsic conformational properties of the diastereomeric backbones. Similar behavior is seen for the diastereomeric peptide pair Ac-L-Val-D-Pro-Gly-L-Leu-NMe(2) vs Ac-L-Val-L-Pro-Gly-L-Leu-NMe(2), and for the analogous depsipeptides with a lactic acid or glycolic acid residue at the third position. Thus, our results show not only that mirror-image Pro-X turns strongly promote beta-hairpin folding, but also that common beta-turns strongly discourage formation of a beta-hairpin with a two-residue loop.