Unusually high alpha-proton acidity of prolyl residues in cyclic peptides

The acidity of the alpha-proton in peptides has an essential role in numerous biochemical reactions and underpins their stereochemical integrity, which is critical to their biological function. We report a detailed kinetic and computational study of the acidity of the alpha-proton in two cyclic peptide systems: diketopiperazine (DKP) and triketopiperazine (TKP). The kinetic acidity (protofugality) of the alpha-protons were determined though hydrogen deuterium exchange studies in aqueous solutions. The acidities of the alpha-proton in prolyl residues were increased by 3-89 fold relative to other amino acid residues (prolyl > glycyl >> alanyl > tyrosyl). Experimental and computational evidence for the stereoelectronic origins of this enhanced prolyl reactivity is presented. TKPs were 10(6)-fold more reactive than their DKP analogues towards deprotonation, which we attribute to the advanced development of aromaticity in the earlier transition state for proton transfer in these cases. A Bronsted linear free energy analysis of the reaction data was conducted to provide estimates of alpha-proton pK(a)s.