Study on physiological role of proline containing peptide

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Summary of research results is the following.1.Synthesis of m orphiceptin(Tyr-Pro-Phe-Pro-NH_2) by dipeptidylaminopeptidase IV(DPIV)Aspergillus oryzae RIB915, isolated from soy sauce koji, was used as the test strain to prepare bran koji, and crude DPIV enzyme solution was obtained through partial purification by salting-out and gel filtration. Conditions for synthesis with this crude enzyme were examined using Tyr-Pro-OEt and Phe-Pro-NH_2 as substrates, and resulted in 40% morphiceptin. This study achieved synthesis of the proline-containing morphiceptin for the first time. In addition, although it has been considered that the protection of the amino group is effective in peptide formation in general, it was shown that peptide formation by DPIV required no such protecting groups.2.Synthesis of Leu-Pro-Tyr-Pro-NH_2 by DPIVWith DPIV that was used in the synthesis of morphiceptin, a two-step reaction including(1)Tyr-Pro-OEt+Arg-NH_2 → Tyr-Pro-Arg-NH_2(2)Leu-Pro-OEt+Tyr-Pro-Arg-NH_2 → Leu … More -Pro-Tyr-Pro-Arg-NH_2to synthesize Leu-Pro-Tyr-Pro-Arg-NH_2 by DPIV was examined in commonly used reaction systems: the aqueous system and the system with addition of organic solvent. Synthesis yield in aqueous system were 31.2% for Tyr-Pro-Arg-NH_2, and 3.3% for Leu-Pro-Tyr-Pro-Arg-NH_2. On the other hand, yields in the system with organic solvent ethylene glycol were 30.5% for Tyr-Pro-Arg-NH_2, and 26.7% for Leu-Pro-Tyr-Pro-Arg-NH_2, showing improvement. This resulted from high synthesis yields for both Tyr-Pro-Arg-NH_2 and Leu-Pro-Tyr-Pro-Arg-NH_2 in the system with addition of ethylene glycol as organic solvent.3.Absorption of opioid peptidesAlthough food-derived dipeptides and tripeptides are known to be absorbed in the small intestine via a transporter, it remains unclear if the pentapeptide Leu-Pro-Tyr-Pro-Arg was also absorbed there. In the in vitro experimental system in rats, the peptide was found in portal blood 26 min after intragastric administration ; LC/MS analysis confirmed that it was Leu-Pro-Tyr-Pro-Arg. Less