Abstract
d-Serine is known to act as an endogenous co-agonist of the N-methyl-d-aspartate receptor in the mammalian brain and is endogenously synthesized from l-serine by a pyridoxal 5′-phosphate-dependent enzyme, serine racemase. Though the soil-living mycetozoa Dictyostelium discoideum possesses no genes homologous to that of NMDA receptor, it contains genes encoding putative proteins relating to the d-serine metabolism, such as serine racemase, d-amino acid oxidase, and d-serine dehydratase. D. discoideum is an attractive target for the elucidation of the unknown functions of d-serine such as a role in cell development. As part of the elucidation of the role of d-serine in D. discoideum, we cloned, overexpressed, and examined the properties of the putative serine racemase exhibiting 46% amino acid sequence similarity with the human enzyme. The enzyme is unique in its stimulation by monovalent cations such as Na+ in addition to Mg2+ and Ca2+, which are well-known activators for the mammalian serine racemase. Mg2+ or Na+ binding caused two- to ninefold enhancement of the rates of both racemization and dehydration. The half-maximal activation concentrations of Mg2+ and Na+ were determined to be 1.2 μM and 2.2 mM, respectively. In the l-serine dehydrase reaction, Mg2+ and Na+ enhanced the k cat value without changing the K m value. Alanine mutation of the residues E207 and D213, which correspond to the Mg2+-binding site of Schizosaccharomyces pombe serine racemase, abolished the Mg2+- and Na+-dependent stimulation. These results suggest that Mg2+ and Na+ share the common metal ion-binding site.
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Abbreviations
- SR:
-
Serine racemase
- DdSR:
-
Serine racemase of Dictyostelium discoideum
- PLP:
-
Pyridoxal 5′-phosphate
- 2,4-DNP:
-
2,4-Dinitrophenyl hydrazine
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Acknowledgments
This work was supported in part by the Program for Promotion of Basic and Applied Researches for Innovation in Bio-oriented Industry (BRAIN) and the Grant-in-Aid for Scientific Research 22380058 (to T. Y.), and the Grant-in-Aid for Research Activity start-up 22880014 (to T. I.) from the Ministry of Education, Culture, Sports, Science, and Technology. We thank Professor Hideko Urushihara of the University of Tsukuba and Japan National Bio Resource Project (NBRP-nenkin) for providing us the EST clone of DdSR.
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T. Ito, H. Murase, and M. Maekawa contributed equally to this work.
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Ito, T., Murase, H., Maekawa, M. et al. Metal ion dependency of serine racemase from Dictyostelium discoideum . Amino Acids 43, 1567–1576 (2012). https://doi.org/10.1007/s00726-012-1232-z
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DOI: https://doi.org/10.1007/s00726-012-1232-z