Synthesis of morphiceptin (Tyr-Pro-Phe-Pro-NH2) by dipeptidyl aminopeptidase IV derived from Aspergillus oryzae

Morphiceptin (Tyr-Pro-Phe-Pro-NH2), tetrapeptide, was synthesized using dipeptidyl aminopeptidase IV (DP IV, EC 3.4.14.5) derived from Aspergillus oryzae RIB 915 as a catalyst. Tyr-Pro-OEt was incubated with Phe-Pro-NH2 in the presence of DP IV under various conditions of temperature, concentrations of ethylene glycol, pH, reaction time, and others. Morphiceptin was obtained at 40% yield under the optimal reaction conditions:  substrate, 4 mM Tyr-Pro-OEt·HCl and 20 mM Phe-Pro-NH2·HCl; enzyme, DP IV, 0.275 nkat; solvent, 60% ethylene glycol containing 20 mM phosphate buffer at pH 7.0; amine, 4.2 mM diisopropylamine at 4 °C for 24 h. Amino group protection was unnecessary for synthesis of morphiceptin by DP IV. Keywords: Morphiceptin; opioid peptide; tetrapeptide; dipeptidyl aminopeptidase IV (DP IV); Koji mold (Aspergillus oryzae); enzymatic synthesis; proline-containing peptide