The presence of an intramolecular hydrogen bond has been proposed to play a key role in the catalysis of amide isomerization by peptidylprolyl isomerases (PPIases), which are highly conserved and ubiquitous rotamase enzymes that catalyze the cis-trans isomerization of proline residues in peptides and proteins. We present herein kinetic and spectroscopic evidence that indicates the existence of an intramolecular hydrogen bond between the prolyl amide nitrogen and the adjacent amidic NH within a five-membered ring (the 5-NN- -N-a hydrogen bond) that is capable of catalyzing proline isomerization by up to 260-fold in model prolyl peptides. Our results provide the first systematic study of intramolecular general-acid-catalyzed amide isomerization.