Enzymatic production of designed peptide

Peptides are expected to be one of the most promising compounds that are beneficial for improving our quality of life. Research on functional peptides has been carried out in various fields, including food science, medicine, and cosmetics; new findings are frequently reported. Oligopeptides such as dipeptides or tripeptides also have unique physiological functions and physical properties that cannot be found in the constitutive amino acids. However, only a few dipeptides, such as l-aspartyl-l-phenylalanine methyl ester (Asp-Phe-OMe, aspartame), as an artificial sweetener and l-alanyl-l-glutamine (Ala-Gln), as a patient infusion, are commercially used, which can be attributed to the lack of an efficient process for production of these oligopeptides. Therefore, the development of an oligopeptide manufacturing process is important for addressing the growing needs of functional peptides. Recently, bacterial enzymes that produce various dipeptides, oligopeptides, or homopoly(oligo) amino acids have been found. l-Amino acid α-ligase (Lal, EC 6.3.2.28) belongs to the ATP-dependent (ADP-forming) carboxylate-amine/thiol ligase superfamily that catalyzes the condensation of unprotected amino acids and is applicable to fermentative production. In this group, ATP and Mg²⁺ are generally required for peptide synthesis, and aminoacyl phosphate is synthesized as the reaction intermediate. Various Lals have been newly identified by in silico searches using a BLAST program and by different approaches including purification of putative Lal from microorganisms producing peptide antibiotics as secondary metabolites. Furthermore, using only an adenylation domain (A-domain) of nonribosomal peptide synthetase (NRPS), various aminoacyl prolines, which are dipeptides containing a proline residue at the C-terminus, or various amide compounds can be synthesized from unprotected amino acids and proline without any additional process. This chapter reviews the current knowledge about these unique enzymes and novel enzymatic production methods of designed peptides.