Amino Acids

, Volume 48, Issue 2, pp 387–402 | Cite as

Distribution and evolution of the serine/aspartate racemase family in invertebrates

  • Kouji Uda
  • Keita Abe
  • Yoko Dehara
  • Kiriko Mizobata
  • Natsumi Sogawa
  • Yuki Akagi
  • Mai Saigan
  • Atanas D. Radkov
  • Luke A. Moe
Original Article


Free d-amino acids have been found in various invertebrate phyla, while amino acid racemase genes have been identified in few species. The purpose of this study is to elucidate the distribution, function, and evolution of amino acid racemases in invertebrate animals. We searched the GenBank databases, and found 11 homologous serine racemase genes from eight species in eight different invertebrate phyla. The cloned genes were identified based on their maximum activity as Acropora millepora (Cnidaria) serine racemase (SerR) and aspartate racemase (AspR), Caenorhabditis elegans (Nematoda) SerR, Capitella teleta (Annelida) SerR, Crassostrea gigas (Mollusca) SerR and AspR, Dugesia japonica (Platyhelminthes) SerR, Milnesium tardigradum (Tardigrada) SerR, Penaeus monodon (Arthropoda) SerR and AspR and Strongylocentrotus purpuratus (Echinodermata) AspR. We found that Acropora, Aplysia, Capitella, Crassostrea and Penaeus had two amino acid racemase paralogous genes and these paralogous genes have evolved independently by gene duplication at their recent ancestral species. The transcriptome analyses using available SRA data and enzyme kinetic data suggested that these paralogous genes are expressed in different tissues and have different functions in vivo. Phylogenetic analyses clearly indicated that animal SerR and AspR are not separated by their particular racemase functions and form a serine/aspartate racemase family cluster. Our results revealed that SerR and AspR are more widely distributed among invertebrates than previously known. Moreover, we propose that the triple serine loop motif at amino acid positions 150–152 may be responsible for the large aspartate racemase activity and the AspR evolution from SerR.


d-Amino acid Serine racemase Aspartate racemase d-Ser d-Asp 



Aspartate racemase


Aplysia californica d-amino acid racemase 1


Expressed sequence tag


Glutamic-oxaloacetic transaminase 1-like 1


O-acetylserine sulfhydrylase


Pyridoxal 5′-phosphate


Reads per kilobase per million reads


Reads per kilobase per million mapped reads


Serine racemase


Sequence read archive


Reverse transcription polymerase chain reaction



This work was supported by a Grant-in-Aid for Scientific Research in Japan to KU (24770068 and 15K07152).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical standard

The article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

726_2015_2092_MOESM1_ESM.pdf (245 kb)
Supplementary material 1 (PDF 245 kb)
726_2015_2092_MOESM2_ESM.pdf (25 kb)
Supplementary material 2 (PDF 25 kb)


  1. Abe H, Yoshikawa N, Sarower MG, Okada S (2005) Physiological function and metabolism of free d-alanine in aquatic animals. Biol Pharm Bull 28(9):1571–1577CrossRefPubMedGoogle Scholar
  2. Abe K, Takahashi S, Muroki Y, Kera Y, Yamada RH (2006) Cloning and expression of the pyridoxal 5′-phosphate-dependent aspartate racemase gene from the bivalve mollusk Scapharca broughtonii and characterization of the recombinant enzyme. J Biochem 139(2):235–244. doi: 10.1093/jb/mvj028 CrossRefPubMedGoogle Scholar
  3. Campbell LI, Rota-Stabelli O, Edgecombe GD, Marchioro T, Longhorn SJ, Telford MJ, Philippe H, Rebecchi L, Peterson KJ, Pisani D (2011) MicroRNAs and phylogenomics resolve the relationships of Tardigrada and suggest that velvet worms are the sister group of Arthropoda. Proc Natl Acad Sci 108(38):15920–15924PubMedCentralCrossRefPubMedGoogle Scholar
  4. Corrigan JJ, Srinivasan N (1966) The occurrence of certain d-amino acids in insects*. Biochemistry 5(4):1185–1190CrossRefPubMedGoogle Scholar
  5. D’Aniello A (2007) d-Aspartic acid: an endogenous amino acid with an important neuroendocrine role. Brain Res Rev 53(2):215–234. doi: 10.1016/j.brainresrev.2006.08.005 CrossRefPubMedGoogle Scholar
  6. Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32(5):1792–1797PubMedCentralCrossRefPubMedGoogle Scholar
  7. Foltyn VN, Bendikov I, De Miranda J, Panizzutti R, Dumin E, Shleper M, Li P, Toney MD, Kartvelishvily E, Wolosker H (2005) Serine racemase modulates intracellular d-serine levels through an alpha, beta-elimination activity. J Biol Chem 280(3):1754–1763. doi: 10.1074/jbc.M405726200 CrossRefPubMedGoogle Scholar
  8. Fujitani Y, Nakajima N, Ishihara K, Oikawa T, Ito K, Sugimoto M (2006) Molecular and biochemical characterization of a serine racemase from Arabidopsis thaliana. Phytochemistry 67(7):668–674. doi: 10.1016/j.phytochem.2006.01.003 CrossRefPubMedGoogle Scholar
  9. Fujitani Y, Horiuchi T, Ito K, Sugimoto M (2007) Serine racemases from barley, Hordeum vulgare L., and other plant species represent a distinct eukaryotic group: gene cloning and recombinant protein characterization. Phytochemistry 68(11):1530–1536. doi: 10.1016/j.phytochem.2007.03.040 CrossRefPubMedGoogle Scholar
  10. Gogami Y, Kobayashi A, Ikeuchi T, Oikawa T (2010) Site-directed mutagenesis of rice serine racemase: evidence that Glu219 and Asp225 mediate the effects of Mg2+ on the activity. Chem Biodivers 7(6):1579–1590. doi: 10.1002/cbdv.200900257 CrossRefPubMedGoogle Scholar
  11. Goto M, Yamauchi T, Kamiya N, Miyahara I, Yoshimura T, Mihara H, Kurihara T, Hirotsu K, Esaki N (2009) Crystal structure of a homolog of mammalian serine racemase from Schizosaccharomyces pombe. J Biol Chem 284(38):25944–25952. doi: 10.1074/jbc.M109.010470 PubMedCentralCrossRefPubMedGoogle Scholar
  12. Hamase K, Morikawa A, Zaitsu K (2002) d-Amino acids in mammals and their diagnostic value. J Chromatogr B 781(1):73–91CrossRefGoogle Scholar
  13. Hashimoto A, Nishikawa T, Oka T, Takahashi K, Hayashi T (1992) Determination of free amino acid enantiomers in rat brain and serum by high-performance liquid chromatography after derivatization with N-tert.-butyloxycarbonyl-l-cysteine and o-phthaldialdehyde. J Chromatogr B Biomed Sci Appl 582(1):41–48CrossRefGoogle Scholar
  14. Hoffman HE, Jiraskova J, Ingr M, Zvelebil M, Konvalinka J (2009) Recombinant human serine racemase: enzymologic characterization and comparison with its mouse ortholog. Protein Expr Purif 63(1):62–67. doi: 10.1016/j.pep.2008.09.003 CrossRefPubMedGoogle Scholar
  15. Hoover DM, Lubkowski J (2002) DNAWorks: an automated method for designing oligonucleotides for PCR-based gene synthesis. Nucleic Acids Res 30(10):e43–e43PubMedCentralCrossRefPubMedGoogle Scholar
  16. Horio M, Ishima T, Fujita Y, Inoue R, Mori H, Hashimoto K (2013) Decreased levels of free d-aspartic acid in the forebrain of serine racemase (Srr) knock-out mice. Neurochem Int 62(6):843–847. doi: 10.1016/j.neuint.2013.02.015 CrossRefPubMedGoogle Scholar
  17. Ito T, Murase H, Maekawa M, Goto M, Hayashi S, Saito H, Maki M, Hemmi H, Yoshimura T (2012) Metal ion dependency of serine racemase from Dictyostelium discoideum. Amino Acids 43(4):1567–1576. doi: 10.1007/s00726-012-1232-z CrossRefPubMedGoogle Scholar
  18. Ito T, Maekawa M, Hayashi S, Goto M, Hemmi H, Yoshimura T (2013) Catalytic mechanism of serine racemase from Dictyostelium discoideum. Amino Acids 44(3):1073–1084. doi: 10.1007/s00726-012-1442-4 CrossRefPubMedGoogle Scholar
  19. Jiraskova-Vanickova J, Ettrich R, Vorlova B, E Hoffman H, Lepsik M, Jansa P, Konvalinka J (2011) Inhibition of human serine racemase, an emerging target for medicinal chemistry. Current Drug Targets 12(7):1037–1055Google Scholar
  20. 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 Anal Technol Biomed Life Sci 879(29):3108–3121. doi: 10.1016/j.jchromb.2011.03.062 CrossRefGoogle Scholar
  21. Kim PM, Duan X, Huang AS, Liu CY, Ming GL, Song H, Snyder SH (2010) Aspartate racemase, generating neuronal d-aspartate, regulates adult neurogenesis. Proc Natl Acad Sci USA 107(7):3175–3179. doi: 10.1073/pnas.0914706107 PubMedCentralCrossRefPubMedGoogle Scholar
  22. Maezawa T, Tanaka H, Nakagawa H, Ono M, Aoki M, Matsumoto M, Ishida T, Horiike K, Kobayashi K (2014) Planarian d-amino acid oxidase is involved in ovarian development during sexual induction. Mech Dev 132:69–78. doi: 10.1016/j.mod.2013.12.003 CrossRefPubMedGoogle Scholar
  23. Matsuda S, Katane M, Maeda K, Kaneko Y, Saitoh Y, Miyamoto T, Sekine M, Homma H (2015) Biosynthesis of d-aspartate in mammals: the rat and human homologs of mouse aspartate racemase are not responsible for the biosynthesis of d-aspartate. Amino Acids. doi: 10.1007/s00726-015-1926-0 PubMedGoogle Scholar
  24. Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B (2008) Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 5(7):621–628CrossRefPubMedGoogle Scholar
  25. Nylander JA, Wilgenbusch JC, Warren DL, Swofford DL (2008) AWTY (are we there yet?): a system for graphical exploration of MCMC convergence in Bayesian phylogenetics. Bioinformatics 24(4):581–583CrossRefPubMedGoogle Scholar
  26. Ohnishi M, Saito M, Wakabayashi S, Ishizuka M, Nishimura K, Nagata Y, Kasai S (2008) Purification and characterization of serine racemase from a hyperthermophilic archaeon, Pyrobaculum islandicum. J Bacteriol 190(4):1359–1365. doi: 10.1128/JB.01184-07 PubMedCentralCrossRefPubMedGoogle Scholar
  27. Okuma E, Fujita E, Amano H, Noda H, Abe H (1995) Distribution of free d-amino acids in the tissues of Crustaceans. Fish Sci 61(1):157–160Google Scholar
  28. Okuma E, Watanabe K, Abe H (1998) Distribution of free d-amino acids in Bivalve Mollusks and the effects of physiological conditions on the levels of d-and l-alanine in the tissues of the hard clam Meretrix lusoria. Fish Sci 64(4):606–611CrossRefGoogle Scholar
  29. Preston R (1987) Occurrence of d-amino acids in higher organisms: a survey of the distribution of d-amino acids in marine invertebrates. Comp Biochem Physiol Part B Comp Biochem 87(1):55–62CrossRefGoogle Scholar
  30. Radkov AD, Moe LA (2014) Bacterial synthesis of d-amino acids. Appl Microbiol Biotechnol 98(12):5363–5374. doi: 10.1007/s00253-014-5726-3 CrossRefPubMedGoogle Scholar
  31. Ronquist F, Huelsenbeck JP (2003) MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19(12):1572–1574CrossRefPubMedGoogle Scholar
  32. Rosenberg H, Ennor A (1961) The occurrence of free d-serine in the earthworm. Biochem J 79(2):424PubMedCentralCrossRefPubMedGoogle Scholar
  33. Saitoh Y, Katane M, Kawata T, Maeda K, Sekine M, Furuchi T, Kobuna H, Sakamoto T, Inoue T, Arai H, Nakagawa Y, Homma H (2012) Spatiotemporal localization of d-amino acid oxidase and d-aspartate oxidases during development in Caenorhabditis elegans. Mol Cell Biol 32(10):1967–1983. doi: 10.1128/MCB.06513-11 PubMedCentralCrossRefPubMedGoogle Scholar
  34. Shibata K, Tarui A, Todoroki N, Kawamoto S, Takahashi S, Kera Y, R-h Yamada (2001) Occurrence of N-methyl-l-aspartate in bivalves and its distribution compared with that of N-methyl-d-aspartate and D, l-aspartate. Comp Biochem Physiol B Biochem Mol Biol 130(4):493–500CrossRefPubMedGoogle Scholar
  35. Shibata K, Watanabe T, Yoshikawa H, Abe K, Takahashi S, Kera Y, Yamada RH (2003) Purification and characterization of aspartate racemase from the bivalve mollusk Scapharca broughtonii. Comp Biochem Physiol B Biochem Mol Biol 134(2):307–314CrossRefPubMedGoogle Scholar
  36. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30(12):2725–2729PubMedCentralCrossRefPubMedGoogle Scholar
  37. Tanaka-Hayashi A, Hayashi S, Inoue R, Ito T, Konno K, Yoshida T, Watanabe M, Yoshimura T, Mori H (2015) Is d-aspartate produced by glutamic-oxaloacetic transaminase-1 like 1 (Got1l1): a putative aspartate racemase? Amino Acids 47(1):79–86. doi: 10.1007/s00726-014-1847-3 PubMedCentralCrossRefPubMedGoogle Scholar
  38. Uda K, Suzuki T (2007) A novel arginine kinase with substrate specificity towards d-arginine. Protein J 26(5):281–291CrossRefPubMedGoogle Scholar
  39. Uo T, Ueda M, Nishiyama T, Yoshimura T, Esaki N (2001) Purification and characterization of alanine racemase from hepatopancreas of black-tiger prawn, Penaeus monodon. J Mol Catal B Enzym 12(1):137–144CrossRefGoogle Scholar
  40. Wang L, Ota N, Romanova EV, Sweedler JV (2011) A novel pyridoxal 5′-phosphate-dependent amino acid racemase in the Aplysia californica central nervous system. J Biol Chem 286(15):13765–13774. doi: 10.1074/jbc.M110.178228 PubMedCentralCrossRefPubMedGoogle Scholar
  41. Wolosker H, Sheth KN, Takahashi M, Mothet J-P, Brady RO, Ferris CD, Snyder SH (1999) Purification of serine racemase: biosynthesis of the neuromodulator d-serine. Proc Natl Acad Sci 96(2):721–725PubMedCentralCrossRefPubMedGoogle Scholar
  42. Wolosker H, D’Aniello A, Snyder S (2000) d-aspartate disposition in neuronal and endocrine tissues: ontogeny, biosynthesis and release. Neuroscience 100(1):183–189CrossRefPubMedGoogle Scholar
  43. Yamauchi T, Goto M, Wu HY, Uo T, Yoshimura T, Mihara H, Kurihara T, Miyahara I, Hirotsu K, Esaki N (2009) Serine racemase with catalytically active lysinoalanyl residue. J Biochem 145(4):421–424. doi: 10.1093/jb/mvp010 CrossRefPubMedGoogle Scholar
  44. Yoshikawa N, Dhomae N, Takio K, Abe H (2002) Purification, properties, and partial amino acid sequences of alanine racemase from the muscle of the black tiger prawn Penaeus monodon. Comp Biochem Physiol B: Biochem Mol Biol 133(3):445–453CrossRefGoogle Scholar
  45. Yoshikawa N, Okada S, Abe H (2009) Molecular characterization of alanine racemase in the kuruma prawn Marsupenaeus japonicus. J Biochem 145(2):249–258. doi: 10.1093/jb/mvn162 CrossRefPubMedGoogle Scholar
  46. Yoshikawa N, Ashida W, Hamase K, Abe H (2011) HPLC determination of the distribution of d-amino acids and effects of ecdysis on alanine racemase activity in kuruma prawn Marsupenaeus japonicus. J Chromatogr B Anal Technol Biomed Life Sci 879(29):3283–3288. doi: 10.1016/j.jchromb.2011.04.026 CrossRefGoogle Scholar
  47. Yuasa HJ, Takubo M, Takahashi A, Hasegawa T, Noma H, Suzuki T (2007) Evolution of vertebrate indoleamine 2, 3-dioxygenases. J Mol Evol 65(6):705–714CrossRefPubMedGoogle Scholar
  48. Zhang G, Fang X, Guo X, Li L, Luo R, Xu F, Yang P, Zhang L, Wang X, Qi H (2012) The oyster genome reveals stress adaptation and complexity of shell formation. Nature 490(7418):49–54CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Wien 2015

Authors and Affiliations

  • Kouji Uda
    • 1
    • 2
  • Keita Abe
    • 1
  • Yoko Dehara
    • 1
  • Kiriko Mizobata
    • 1
  • Natsumi Sogawa
    • 1
  • Yuki Akagi
    • 1
  • Mai Saigan
    • 1
  • Atanas D. Radkov
    • 2
  • Luke A. Moe
    • 2
  1. 1.Laboratory of Biochemistry, Faculty of ScienceKochi UniversityKochiJapan
  2. 2.Department of Plant and Soil Sciences, 311 Plant Science BuildingUniversity of KentuckyLexingtonUSA

Personalised recommendations