Synthesis of d-Amino Acid-Containing Dipeptides Using the Adenylation Domains of Nonribosomal Peptide Synthetase.

Abstract

Recent papers have reported dipeptides containing d-amino acids to have novel effects that cannot be observed with ll-dipeptides, and such dipeptides are expected to be novel functional compounds for pharmaceuticals and food additives. Although the functions of d-amino acid-containing dipeptides are gaining more attention, there are few reports on the synthetic enzymes that can accept d-amino acids as substrates, and synthetic methods for d-amino acid-containing dipeptides have not yet been constructed. Previously, we developed a chemoenzymatic system for amide synthesis that comprised enzymatic activation and a subsequent nucleophilic substitution reaction. In this study, we demonstrated the application of the system for d-amino acid-containing-dipeptide synthesis. We chose six adenylation domains as targets according to our newly constructed hypothesis, i.e., an adenylation domain located upstream from the epimerization domain may activate d-amino acid as well as l-amino acid. We successfully synthesized over 40 kinds of d-amino acid-containing dipeptides, including ld-, dl-, and dd-dipeptides, using only two adenylation domains, TycA-A from tyrocidine synthetase and BacB2-A from bacitracin synthetase. Furthermore, this study offered the possibility that the epimerization domain could be a clue to the activity of the adenylation domains toward d-amino acid. This paper provides additional information regarding d-amino acid-containing-dipeptide synthesis through the combination of enzymatic adenylation and chemical nucleophilic reaction, and this system will be a useful tool for dipeptide synthesis.IMPORTANCE Because almost all amino acids in nature are l-amino acids, the functioning of d-amino acids has received little attention. Thus, there is little information available on the activity of enzymes toward d-amino acids or synthetic methods for d-amino acid-containing dipeptides. Recently, d-amino acids and d-amino acid-containing peptides have attracted attention as novel functional compounds, and d-amino acid-activating enzymes and synthetic methods are required for the development of the d-amino acid-containing-peptide industry. This study provides additional knowledge regarding d-amino acid-activating enzymes and proposes a unique synthetic method for d-amino acid-containing peptides, including ld-, dl-, and dd-dipeptides.