Structure-based modification of D-alanine-D-alanine ligase from Thermotoga maritima ATCC 43589 for depsipeptide synthesis

Tomoki Nakagawa, Ryoko Satake, Masaru Sato, Kuniki Kino*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Depsipeptides are peptide-like polymers consisting of amino acids and hydroxy acids, and are expected to be new functional materials for drug-delivery systems and polymer science. In our previous study, D-alanyl-D-lactate, a type of depsipeptide, was enzymatically synthesized using D-alanine-D-alanine ligase from Thermotoga maritima ATCC 43589 (TmDdl) by Y207F substitution. Thereafter, in this study, further mutagenesis was introduced, based on structural comparison between TmDdl and a well-characterized D-alanine-D-alanine ligase from Escherichia coli. The S137A/Y207F mutant showed higher D-alanyl-D-lactate and lower D-alanyl-D-alanine synthesizing activity than the Y207F mutant. This suggests that substitution at the S137 residue contributes to product selectivity. Saturated mutagenesis on S137 revealed that the S137G/Y207F mutant showed the highest D-alanyl-D-lactate synthesizing activity. Moreover, the mutant showed broad substrate specificity toward D-amino acid and recognized D-lactate and D,L-isoserine as substrates. On the basis of these characteristics, various depsipeptides can be produced using S137G/Y207F-replaced TmDdl.

Original languageEnglish
Pages (from-to)700-704
Number of pages5
JournalBioscience, Biotechnology and Biochemistry
Issue number4
Publication statusPublished - 2011


  • D-alanine-D-alanine ligase
  • Depsipeptide
  • Dipeptide
  • Enzymatic peptide synthesis
  • Thermotoga maritima

ASJC Scopus subject areas

  • Biotechnology
  • Analytical Chemistry
  • Biochemistry
  • Applied Microbiology and Biotechnology
  • Molecular Biology
  • Organic Chemistry


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