1. Structure-function relationships in human d-aspartate oxidase: characterisation of variants corresponding to known single nucleotide polymorphisms
- Author
-
Yasuaki Saitoh, Kazuki Nakayama, Masumi Katane, Hiroshi Homma, Masae Sekine, Ryo Kanazawa, Risa Kobayashi, Tetsuya Miyamoto, and Megumi Oishi
- Subjects
0301 basic medicine ,D-aspartate oxidase ,Models, Molecular ,D-Aspartate Oxidase ,endocrine system diseases ,Protein Conformation ,Biophysics ,Flavoprotein ,Biology ,Degradative enzyme ,Transfection ,Biochemistry ,Polymorphism, Single Nucleotide ,Receptors, N-Methyl-D-Aspartate ,Cofactor ,Analytical Chemistry ,Substrate Specificity ,03 medical and health sciences ,chemistry.chemical_compound ,Structure-Activity Relationship ,Cell Line, Tumor ,Excitatory Amino Acid Agonists ,Animals ,Humans ,Pituitary Neoplasms ,Amino Acids ,Molecular Biology ,chemistry.chemical_classification ,Flavin adenine dinucleotide ,Oxidase test ,Aspartic Acid ,Stereoisomerism ,Molecular biology ,Enzyme assay ,Recombinant Proteins ,Amino acid ,Rats ,030104 developmental biology ,chemistry ,Amino Acid Substitution ,biology.protein ,Flavin-Adenine Dinucleotide ,Mutagenesis, Site-Directed ,Excitatory Amino Acid Antagonists ,Protein Binding - Abstract
d-Aspartate oxidase (DDO) is a degradative enzyme that is stereospecific for the acidic amino acid d-aspartate, an endogenous agonist of the N-methyl-d-aspartate (NMDA) receptor. Dysregulation of NMDA receptor-mediated neurotransmission has been implicated in the onset of various neuropsychiatric disorders including schizophrenia and in chronic pain. Thus, appropriate regulation of the amount of d-aspartate is believed to be important for maintaining proper neural activity in the nervous system. Herein, the effects of the non-synonymous single nucleotide polymorphisms (SNPs) R216Q and S308N on several properties of human DDO were examined. Analysis of the purified recombinant enzyme showed that the R216Q and S308N substitutions reduce enzyme activity towards acidic d-amino acids, decrease the binding affinity for the coenzyme flavin adenine dinucleotide and decrease the temperature stability. Consistent with these findings, further experiments using cultured mammalian cells revealed elevated d-aspartate in cultures of R216Q and S308N cells compared with cells expressing wild-type DDO. Furthermore, accumulation of several amino acids other than d-aspartate also differed between these cultures. Thus, expression of DDO genes carrying the R216Q or S308N SNP substitutions may increase the d-aspartate content in humans and alter homeostasis of several other amino acids. This work may aid in understanding the correlation between DDO activity and the risk of onset of NMDA receptor-related diseases.
- Published
- 2017