Enzymatic synthesis of Z-aspartame in liquefied amino acid substrates

In this paper, we designed and validated a new media for biotransformation aided by ionic liquefied amino acid substrates. Thermolysin-catalyzed synthesis of Z-aspartame (N-carbobenzoxy-l-aspartame) in which ionic liquids play dual roles as both substrate and reaction medium was conducted. If the protease can retain catalytic activity in this new liquefied amino acid substrate/media, the effective substrate concentration can be maximized, leading to theoretically high yields. In fact, we observed a 34-fold enhancement in the catalytic activity compared with that obtained in a previous report. The yield of Z-aspartame reached at approximately 660 mM at 24 h with 50 mg mL−1 of the enzyme, thus submolar range productivity was achieved. The results suggest the successful construction of a high productivity reaction system for peptide synthesis.

► A new media for biotransformation aided by ionic liquefied amino acid substrates was designed. ► The amino acid ionic liquids play dual roles as substrates and reaction media. ► In this reaction media, effective peptide substrate concentration can be maximized. ► Thermolysin exhibited the catalytic activity in the presence of 10 vol% buffered water. ► The submolar range productivity in protease-catalyzed peptide synthesis was achieved.