Characterization of an α-amino-ɛ-caprolactam racemase with broad substrate specificity from Citreicella sp. SE45

  • Wisarut Payoungkiattikun
  • Seiji Okazaki
  • Atsutoshi Ina
  • Aran H-Kittikun
  • Yasuhisa Asano
Biocatalysis - Original Paper


α-Amino-ε-caprolactam (ACL) racemizing activity was detected in a putative dialkylglycine decarboxylase (EC from Citreicella sp. SE45. The encoding gene of the enzyme was cloned and transformed in Escherichia coli BL21 (DE3). The molecular mass of the enzyme was shown to be 47.4 kDa on SDS–polyacrylamide gel electrophoresis. The enzymatic properties including pH and thermal optimum and stabilities were determined. This enzyme acted on a broad range of amino acid amides, particularly unbranched amino acid amides including l-alanine amide and l-serine amide with a specific activity of 17.5 and 21.6 U/mg, respectively. The K m and V max values for d- and l-ACL were 5.3 and 2.17 mM, and 769 and 558 μmol/ protein, respectively. Moreover, the turn over number (K cat) and catalytic efficiency (K cat/K m ) of purified ACL racemase from Citreicella sp. SE45 using l-ACL as a substrate were 465 S−1 and 214 S−1mM−1, respectively. The new ACL racemase from Citreicella sp. SE45 has a potential to be used as the biocatalytic application.


ACL racemase PLP-dependent enzyme d-amino acids Enzyme characterization Citreicella sp. SE45 



This work was financially supported by the Thailand Research Fund through the Royal Golden Jubilee Ph.D. Program (Grant No. PHD/099/2551) and the Japan Student Services Organization (JASSO). This research was partly supported by Grant-in-Aids for Scientific Research (A) (23248015) and (B) (26292041) to Y. Asano from the Japan Society for the Promotion of Science (JSPS). This work was also partly supported by the Exploratory Research for Advanced Technology (ERATO) Asano Active Enzyme Molecule Project of Japan Science and Technology Agency (JST). The author greatly appreciates Asst. Prof. Ken-ichi Fuhshuku for the preparation of chiral ACLs and enantiomerically pure amino acid amides. This work was performed at Toyama Prefectural University.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no completing interests.

Ethical approval

This study does not contain any experiment with human participants or animals performed by any of the authors.

Supplementary material

10295_2016_1825_MOESM1_ESM.docx (270 kb)
Supplementary material 1 (DOCX 270 kb)


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Copyright information

© Society for Industrial Microbiology and Biotechnology 2016

Authors and Affiliations

  1. 1.Department of Industrial Biotechnology, Faculty of Agro-IndustryPrince of Songkla UniversityHat-YaiThailand
  2. 2.Department of Biotechnology and Biotechnology Research CenterToyama Prefectural UniversityImizuJapan
  3. 3.Asano Active Enzyme Molecule Project, ERATO, JSTImizuJapan

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