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Efficient synthesis of d-branched-chain amino acids and their labeled compounds with stable isotopes using d-amino acid dehydrogenase

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Abstract

d-Branched-chain amino acids (d-BCAAs) such as d-leucine, d-isoleucine, and d-valine are known to be peptide antibiotic intermediates and to exhibit a variety of bioactivities. Consequently, much effort is going into achieving simple stereospecific synthesis of d-BCAAs, especially analogs labeled with stable isotopes. Up to now, however, no effective method has been reported. Here, we report the establishment of an efficient system for enantioselective synthesis of d-BCAAs and production of d-BCAAs labeled with stable isotopes. This system is based on two thermostable enzymes: d-amino acid dehydrogenase, catalyzing NADPH-dependent enantioselective amination of 2-oxo acids to produce the corresponding d-amino acids, and glucose dehydrogenase, catalyzing NADPH regeneration from NADP+ and d-glucose. After incubation with the enzymes for 2 h at 65°C and pH 10.5, 2-oxo-4-methylvaleric acid was converted to d-leucine with an excellent yield (>99 %) and optical purity (>99 %). Using this system, we produced five different d-BCAAs labeled with stable isotopes: d-[1-13C,15N]leucine, d-[1-13C]leucine, d-[15N]leucine, d-[15N]isoleucine, and d-[15N]valine. The structure of each labeled d-amino acid was confirmed using time-of-flight mass spectrometry and nuclear magnetic resonance analysis. These analyses confirmed that the developed system was highly useful for production of d-BCAAs labeled with stable isotopes, making this the first reported enzymatic production of d-BCAAs labeled with stable isotopes. Our findings facilitate tracer studies investigating d-BCAAs and their derivatives.

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References

  • Akita H, Fujino Y, Doi K, Ohshima T (2011) Highly stable meso-diaminopimelate dehydrogenase from an Ureibacillus thermosphaericus strain A1 isolated from a Japanese compost: purification, characterization and sequencing. AMB Express 1:43

    Article  PubMed Central  PubMed  Google Scholar 

  • Akita H, Doi K, Kawarabayasi Y, Ohshima T (2012) Creation of a thermostable NADP+-dependent d-amino acid dehydrogenase from Ureibacillus thermosphaericus strain A1 meso-diaminopimelate dehydrogenase by site-directed mutagenesis. Biotechnol Lett 34:1693–1699, Erratum to this report can be found in Biotechnol Lett 34:1701–1702 (2012)

    Article  CAS  PubMed  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Chaohui Y, Riqiang F, Jianzhi H, Lei H, Shangwu D (1993) Carbon-13 chemical shift anisotropies of solid amino acids. Magn Reson Chem 31:699–704

    Article  Google Scholar 

  • Chiriac M, Lupan I, Bucurenci N, Popescu O, Palibroda N (2007) Stereoselective synthesis of l-[15N] amino acids with glucose dehydrogenase and galactose mutarotase as NADH regenerating system. J Label Compd Radiopharm 51:171–174

    Article  Google Scholar 

  • Doveston RG, Steendam R, Jones S, Taylor RJ (2012) Total synthesis of an oxepine natural product, (±)-janoxepin. Org Lett 14:1122–1125

    Article  CAS  PubMed  Google Scholar 

  • Ellis AV, Wilson MA (2002) Carbon exchange in hot alkaline degradation of glucose. J Org Chem 24:8469–8474

    Article  Google Scholar 

  • Friedman M (2010) Origin, microbiology, nutrition, and pharmacology of d-amino acids. Chem Biodivers 7:1491–1530

    Article  CAS  PubMed  Google Scholar 

  • Friedman M, Levin CE (2012) Nutritional and medicinal aspects of d-amino acids. Amino Acids 42:1553–1582

    Article  CAS  PubMed  Google Scholar 

  • Heine W, Wutzke K, Drescher U (1983) d-Amino acid utilization in infants measured with the 15N-tracer technique. Clin Nutr 2:31–35

    Article  CAS  PubMed  Google Scholar 

  • Jones JG, Cottam GL, Miller BC, Sherry AD, Malloy CR (1994) A method for obtaining 13C isotopomer populations in 13C-enriched glucose. Anal Biochem 217:148–152

    Article  CAS  PubMed  Google Scholar 

  • Katane M, Homma H (2011) d-Aspartate—an important bioactive substance in mammals: a review from an analytical and biological point of view. J Chromatogr B 879:3108–3121

    Article  CAS  Google Scholar 

  • Kawagishi H, Hamajima K, Takanami R, Nakamura T, Sato Y, Akiyama Y, Sano M, Tanaka O (2004) Growth promotion of mycelia of the matsutake mushroom Tricholoma matsutake by d-isoleucine. Biosci Biotechnol Biochem 68:2405–2407

    Article  CAS  PubMed  Google Scholar 

  • Kazimierczuk K, Misiak M, Zawadzka A, Koźmiński W (2007) Progress in structural studies of proteins by NMR spectroscopy. Polimery 52:736–744

    CAS  Google Scholar 

  • Kolodkin-Gal I, Romero D, Cao S, Clardy J, Kolter R, Losick R (2010) d-Amino acids trigger biofilm disassembly. Science 328:627–629

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lankiewicz L, Nyassé B, Fransson B, Grehn L, Ragnarsson U (1994) Synthesis of amino acid derivatives substituted in the backbone with stable isotopes for application in peptide synthesis. J Chem Soc Perkin Trans 1:2503–2510

    Article  Google Scholar 

  • Martínez-Rodríguez S, Martínez-Gómez AI, Rodríguez-Vico F, Clemente–Jiménez JM, Las Heras-Vázquez FJ (2010) Natural occurrence and industrial applications of d-amino acids: an overview. Chem Biodivers 7:1531–1548

    Article  PubMed  Google Scholar 

  • Nimura N, Kinoshita T (1986) o-Phthalaldehyde-N-acetyl-l-cysteine as a chiral derivatization reagent for liquid chromatographic optical resolution of amino acid ernantiomers and its application to conventional amino acid analysis. J Chromatogr A 352:169–177

    Article  CAS  Google Scholar 

  • Nishikawa T (2011) Analysis of free d-serine in mammals and its biological relevance. J Chromatogr B 879:3169–3183

    Article  CAS  Google Scholar 

  • Ogrel A, Shvets VI, Kaptein B, Broxterman QB, Jan R (2000) Synthesis of 15N-labelled d-isovaline and α-aminoisobutyric acid. Eur J Org Chem 2000:857–859

    Article  Google Scholar 

  • Ohmori T, Mutaguchi Y, Yoshikawa S, Doi K, Ohshima T (2011) Amino acid components of lees in salmon fish sauce are tyrosine and phenylalanine. J Biosci Bioeng 112:256–258

    Article  CAS  PubMed  Google Scholar 

  • Ohshima T, Ito Y, Sakuraba H, Goda S, Kawarabayasi Y (2003) The Sulfolobus tokodaii gene ST1704 codes highly thermostable glucose dehydrogenase. J Mol Catal B Enzym 23:281–289

    Article  CAS  Google Scholar 

  • Panayiotis A, Karl O, Sheo BS (1984) Synthesis of (2RS)-[3-13C]- and (2RS)-[3-14C]leucine. J Label Compd Radiopharm 21:393–400

    Article  Google Scholar 

  • Polach KJ, Shah SA, Laluppa JC, Lemaster DM (1993) Racemic synthesis and enantiomeric conversion of [1-13C] valine. J Label Compd Radiopharm 33:809–815

    Article  CAS  Google Scholar 

  • Reid CM, Sutherland A (2009) 11 Synthesis of isotopically labeled α-amino acids. In: Hughes AB (ed) Amino acids, peptides and proteins in organic chemistry, volume 1, origins and synthesis of amino acids. Wiley, New Jersey, pp 473–493

    Google Scholar 

  • Sasaki T, Takahashi S, Uchida K, Funayama S, Kainosho M, Nakagawa A (2006) Biosynthesis of quinolactacin A, a TNF production inhibitor. J Antibiot (Tokyo) 59:418–427

    Article  CAS  Google Scholar 

  • Scapin G, Reddy SG, Blanchard JS (1996) Three-dimensional structure of meso-diaminopimelic acid dehydrogenase from Corynebacterium glutamicum. Biochemistry 35:13540–13551

    Article  CAS  PubMed  Google Scholar 

  • Tempelaars MH, Rodrigues S, Abee T (2011) Comparative analysis of antimicrobial activities of valinomycin and cereulide, the Bacillus cereus emetic toxin. Appl Environ Microbiol 77:2755–2762

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Tiainen M, Maaheimo H, Soininen P, Laatikainen R (2010) 13C isotope effects on 1H chemical shifts: NMR spectral analysis of 13C-labelled d-glucose and some 13C-labelled amino acids. Magn Reson Chem 48:117–122

    Article  CAS  PubMed  Google Scholar 

  • Velkov T, Thompson PE, Nation RL, Li J (2010) Structure–activity relationships of polymyxin antibiotics. J Med Chem 53:1898–1916

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Wałaszek Z, Horton D, Ekiel I (1982) Conformational studies on aldonolactones by N.M.R. spectroscopy. Conformations of d-glucono-1,5-lactone and d-mannono-1,5-lactone in solution. Carbohydr Res 106:193–201

    Article  Google Scholar 

  • Wang W, Liu G, Yamashita K, Manabe M, Kodama H (2004) Characteristics of prolinase against various iminodipeptides in erythrocyte lysates from a normal human and a patient with prolidase deficiency. Clin Chem Lab Med 42:1102–1108

    Article  CAS  PubMed  Google Scholar 

  • Yuan S, Ajami AM (1985) Synthesis of l-[1-13C]isoleucine via amidocarbonylation. J Label Compd Radiopharm 22:1309–1314

    Article  CAS  Google Scholar 

  • Yuan S, Foos J (1981) Synthesis of (S)-leucine-13C3 and its metabolites. J Label Compd Radiopharm 18:563–569

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank Professor Seiki Kuramitsu and Ms. Keiko Ideta for TOF-MS and NMR analyses, respectively. This work was supported by a grant for Promotion of Basic Research Activities for Innovate Bioscience from the Bio-oriented Technology Research Advancement Institution (BRAIN) and Geo Biotechnology Development Organization.

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Authors and Affiliations

  1. Applied Molecular Microbiology and Biomass Chemistry, Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, 812-8581, Japan

    Hironaga Akita

  2. Microbial Genetic Division, Institute of Genetic Resources, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, 812-8581, Japan

    Katsumi Doi & Toshihisa Ohshima

  3. Functional Genomics of Extremophiles, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, 812-8581, Japan

    Hirokazu Suzuki

Authors
  1. Hironaga Akita
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  2. Hirokazu Suzuki
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  3. Katsumi Doi
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  4. Toshihisa Ohshima
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Correspondence to Toshihisa Ohshima.

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Akita, H., Suzuki, H., Doi, K. et al. Efficient synthesis of d-branched-chain amino acids and their labeled compounds with stable isotopes using d-amino acid dehydrogenase. Appl Microbiol Biotechnol 98, 1135–1143 (2014). https://doi.org/10.1007/s00253-013-4902-1

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  • DOI: https://doi.org/10.1007/s00253-013-4902-1

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