Characterization of Tyr-Leu-Gly, a novel anxiolytic-like peptide released from bovine αS-casein

We found previously that dipeptide YL exhibits orally active anxiolytic activity comparable to diazepam. The YL sequence is often observed in the primary structure of natural food proteins. In the present study, we investigated whether YL and YL analogues are released from bovine (S)-casein by gastrointestinal proteases. YLG, corresponding to (S1)-casein (aa 91-93), was more effectively released from (S)-casein than YL by pepsin-pancreatin digestion, mimicking gastrointestinal enzymatic conditions. Using the synthetic model peptide, we determined that trypsin cleaved the N terminus of YLG, and elastase and carboxypeptidase contributed to cleave the C-terminus. YLG exhibited orally active anxiolytic-like activity in the elevated plus maze and open-field tests in mice. The anxiolytic-like activity of YLG was inhibited by WAY100135, SCH23390 or bicuculline, antagonists of serotonin 5-HT1A, dopamine D-1, and GABA(A) receptors, respectively; however, YLG had no affinity for these receptors. The pepsin-pancreatin digest of (S)-Casein also exhibited anxiolytic-like activity. Meanwhile, anxiolytic-like activity of -casozepine, an (S1)-casein-derived decapeptide with YL sequence in the N terminus, was blocked by WAY100135, SCH23390, or bicuculline, equally to YLG and YL; however, it was not detected in the pepsin-pancreatic digest. Taken together, we found that YLG is released after pepsin-pancreatic digestion of (S)-casein and exhibits potent anxiolytic-like activity via activation of serotonin, dopamine, and the GABA receptor system.Mizushige, T., Sawashi, Y., Yamada, A., Kanamoto, R., Ohinata, K. Characterization of Tyr-Leu-Gly, a novel anxiolytic-like peptide released from bovine (S)-casein.