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The Editor’s pick selection of the most intriguing papers is highlighted in yellow.



  • Effects of D-amino acids on sleep in Drosophila

Hiroyuki Nakagawa, Shin Nakane, Gosuke Ban, Jun Tomita, Kazuhiko Kume
Biochemical and Biophysical Research Communications, Volume 589, 2022, 180-185,


Here, a comprehensively investigation about the effects of D-amino acids fed in the diet on the sleep of Drosophila melanogaster was performed. Both D-Ser and D-Gln induced a significant increase in sleep amount (D-Ser showed the highest effect at 1% of the food). D-Ser activates NMDA type glutamate receptor: accordingly, it was not able to increase the sleep of the NR1 hypomorphic mutant flies. On the other hand, hypomorphic mutants of D-amino acid oxidase (Daao1), which inactivation yields an increase in D-Ser levels in the brain, showed increase in sleep. These results suggest that D-Ser, by activating NMDA receptors in the brain, increases sleep, and that D-Ser physiologically regulates sleep.



  • Serine racemase expression by striatal neurons


Takagi S, Puhl MD, Anderson T, Balu DT, Coyle JT. 
Cell Mol Neurobiol. (2022) 42(1):279-289. doi: 10.1007/s10571-020-00880-9.

The localization of D-Ser and serine racemase (SR) was investigated in the mouse striatum, as well as the effects of genetically silencing SR expression in GABAergic interneurons (iSR-/-). iSR-/- mice had substantially reduced SR expression, almost exclusively in striatum, while D-Ser level was marginally reduced. In wild-type mice, the GABAergic medium spiny neurons receiving dopaminergic inputs in striatum robustly and uniformly express SR. In behavioral tests, iSR-/- mice showed a blunted response to the hedonic and stimulant effects of cocaine, without affecting anxiety-related behaviors. Because the cocaine effects have been reported in the constitutive SR-/- mice, the restriction of the blunted response to cocaine to iSR-/- mice reinforces the conclusion that in striatal GABAergic neurons D-Ser plays a main role in mediating dopaminergic stimulant effects. In conclusion, SR in striatal GABAergic neurons synthesizes D-Ser, not as a glutamatergic co-transmitter but as an autocrine signal; the GABAergic neurons control the excitability of their NMDARs by controlling the levels of the co-agonist D-Ser.





  • Effects of sodium benzoate, a D-amino acid oxidase inhibitor, on perceived stress and cognitive function among patients with late-life depression: A randomized, double-blind, sertraline- and placebo-controlled trial 

Chieh-Hsin Lin, Shi-Heng Wang, Hsien-Yuan Lane
International Journal of Neuropsychopharmacology, pyac006,

This work aims at comparing sodium benzoate (a well-known DAAO inhibitor and an indirect NMDAR enhancer), sertraline (a selective serotonin reuptake inhibitor), and placebo in the treatment of late-life depression. A total of 117 patients with major depressive disorder aged >55 years received 8-week treatment of 250-1500 mg/day of sodium benzoate or 25-150 mg/day of sertraline. Compared to placebo, sodium benzoate but not sertraline substantially improved Perceived Stress Scale (PSS) scores and cognitive function, while sertraline, but not benzoate, significantly reduced self-report Geriatric Depression Scale (GDS) scores. Sertraline treatment raised low-density lipoprotein levels. Benzoate-treated patients were less likely to drop out than sertraline- or placebo-recipients. The authors concluded that sertraline can reduce subjective depressive symptoms, while benzoate can decrease perceived stress, improve cognitive function, and enhance treatment adherence in late-life depression patients.



  • Efficacy and safety of add-on sodium benzoate, a D-amino acid oxidase inhibitor, in treatment of schizophrenia: A systematic review and meta-analysis

Seetharam JC, Maiti R, Mishra A, Mishra BR. 
Asian J Psychiatr. (2021) 68:102947. doi: 10.1016/j.ajp.2021.102947


This review focuses on the role of sodium benzoate (a D-amino acid oxidase inhibitor) in schizophrenia. Add-on sodium benzoate significantly improved positive symptoms of schizophrenia but had no important favourable effect on negative symptoms, general psychopathology, and total PANSS score compared to control. Indeed, there was no significant improvement in GAF, CGI, cognitive function and quality of life. Extrapyramidal symptoms were significantly higher in the group treated with sodium benzoate than in the control group, with no significant difference in other adverse effects. The authors conclude that sodium benzoate can improve the positive symptoms of schizophrenia without any beneficial effect on other symptomatology, cognition, quality of life and functioning.



  • Phenothiazine, anthraquinone and related tricyclic derivatives as inhibitors of D-amino acid oxidase

Roslyn Lefin, Anél Petzer, Stephanus  J. Cloete, Jacobus P. Petzer
Results in Chemistry (2022) Volume 4, 100278

A recent study reported that phenothiazine, anthraquinone and related tricyclic derivatives are good potency inhibitors of the monoamine oxidase enzymes. This paper investigated whether these compounds may also inhibit D-amino acid oxidase (DAAO). Seven compounds exhibit good potency inhibition of DAAO with IC50 values in the 0.3-6.3 µM range), values comparable to that of the reference inhibitor 3-methylpyrazole-5-carboxylic acid (IC50 = 1.3 µM). The most potent inhibitors are 2-chloro-7-methoxy-10H-phenothiazine (IC50 = 0.31 µM), 2-chlorophenothiazine (IC50 = 0.33 µM) and quinizarin (IC50 = 0.37 µM).



  • D-Amino acids and D-amino acid-containing peptides: potential disease biomarkers and therapeutic targets? 

Abdulbagi M, Wang L, Siddig O, Di B, Li B. 
Biomolecules (2021) 18;11(11):1716. doi: 10.3390/biom11111716

Significant levels of free D-amino acids were identified in several diseases and various D-amino acid-containing peptides (DAACPs) were isolated from patients with cataracts, Alzheimer’s and other pathologies. Furthermore, recent advances in analytical techniques led to improvement in the discovery and analysis of DAACPs and D-amino acids This review focuses on the presence of DAACPs and free D-amino acids and on their links with disease development and progress.



  • Association of the D-amino acid oxidase gene with methadone dose in heroin dependent patients under methadone maintenance treatment.

Liu TH, Tsou HH, Chung RH, Liu SC, Wang SC, Kuo HW, Fang CP, Chen ACH, Liu YL.
J Hum Genet. (2022) doi: 10.1038/s10038-021-01008-7

Methadone is a synthetic opioid used for the maintenance treatment (MMT) of heroin dependence (it binds to the μ-opioid receptor) and it is also an N-methyl-D-aspartate (NMDA) receptor antagonist. The role of NMDA receptor in the regulatory mechanisms of methadone dosage in heroin dependent patients is unclear. The hypothesis that genetic polymorphisms in the D-amino acid oxidase (DAAO) encoding gene are associated with methadone treatment dose and responses was studied focusing on four single nucleotide polymorphisms in DAAO: SNPs were genotyped in 344 MMT patients. The SNP rs55944529 reveals a modest but significant association with the methadone dosage in the recessive model of analysis and plasma concentrations in MMT patients. It is also associated with the methadone adverse reactions of dry mouth, difficulty with urination in the dominant model, and the withdrawal symptoms of yawning and gooseflesh skin in the recessive model. This study suggests a role of NMDA receptors, possibly via the DAAO genetic variants, in the methadone dose and some adverse reactions in MMT patients.





  • Determination of amino acid content and its enantiomeric composition in honey samples from Mendoza, Argentina

Pamela Y. Quintas, Sonia Keunchkarian, Lilian Romero, Brenda V. Canizo, Rodolfo G. Wuilloud, Cecilia Castell 
J Food Processing and Preservation (2021) Volume 45, Issue 12, e15966

HPLC and chiral GC coupled to MS were used to investigate the amino acid content of honeys from Mendoza. Proline was the most abundant amino acid, followed by phenylalanine, while D-Pro level was lower than D-Phe one; D-Ala, D-Val, D-Glu, D-Leu, and D-Ile were also present in most samples. The authors propose that certain D-AAs can occur naturally in this honey as a consequence of the Maillard reaction.





  • Single-cell-based screening and engineering of D-amino acid amidohydrolases using artificial amidophenol substrates and microbial biosensors

Yeom SJ, Kwon KK, An JU, Park SH, Lee JY, Rha E, Lee H, Kim H, Lee DH, Lee SG. 
J Agric Food Chem. (2022) doi: 10.1021/acs.jafc.1c05834

This paper focuses on a transcriptional factor-based screening strategy for the rapid screening of D-stereospecific amino acid amidase via an enzyme-specific amidophenol substrate. In details, D-threonine amidophenyl derivative was used to produce 2-aminophenol, acting as a putative enzyme indicator in the presence of D-threonine amidases. By using this approach, several Bacillus strains were demonstrated to produce amidase: putative amidase gene was cloned and used for rapid directed evolution. The effect of F119A substitution was thus reported to significantly improve the catalytic activity on D-Ala, D-Thr, and D-Glu.



  • Discovery of a new flavin N5-adduct in a tyrosine to phenylalanine variant of D-arginine dehydrogenase

Iyer A, Reis RAG, Agniswamy J, Weber IT, Gadda G. 
Arch Biochem Biophys. (2022) 715:109100. doi: 10.1016/

D-Arg dehydrogenase from Pseudomonas aeruginosa (PaDADH) catalyzes the flavin-dependent oxidation of D-Arg and other D-amino acids. Here, the group of Giovanni Gadda reports on the crystal structure at 1.29 Å resolution for PaDADH-Y249F in complex with D-Arg. Electron density maps and mass spectrometric analysis highlighted the presence of two modified flavin molecules, i.e. N5-(4-guanidino-oxobutyl)-FAD and 6-OH-FAD. The versatility of the reduced flavin is apparent in the PaDADH-Y249F structure and is evidenced by the multiple functions it can perform in the same active site.



  • Regulation of D-aspartate oxidase gene expression by pyruvate metabolism in the yeast Cryptococcus humicola

Imanishi D, Zaitsu S, Takahashi S. 
Microorganisms (2021) 27;9(12):2444. doi: 10.3390/microorganisms9122444

In the yeast Cryptococcus humicola strain UJ1, the enzyme D-aspartate oxidase (ChDDO) is essential for D-Asp use and is expressed only in the presence of D-Asp, while the enzyme pyruvate carboxylase (Pyc, converting pyruvate into oxaloacetate) is involved in the import and activation of specific peroxisomal flavoenzymes. PYC gene disruption (∆Chpyc1) resulted in growth retardation on glucose and NH4Cl medium: the growth was restored by adding oxaloacetate from L-Asp while D-Asp did not prevent growth retardation because of a decrease in ChDDO gene expression. Pyruvate significantly decreased ChDDO gene transcription in the ∆Chpyc1 strain but increased the same in the wild-type strain, although the intracellular pyruvate level was similar in both strains. The authors conclude that ChDDO gene expression might be regulated by both pyruvate metabolism and D-Asp.





  • Transient potentiometry based D-serine sensor using engineered D-amino acid oxidase showing quasi-direct electron transfer property

Shouhei Takamatsu, Inyoung Lee, Jinhee Lee, Ryutaro Asano, Wakako Tsugawa, Kazunori Ikebukuro, Jeffrey E. Dick, Koji Sode
Biosensors and Bioelectronics (2022) Volume 200, 113927, ISSN 0956-5663,

This paper reports on a novel sensing principle for D-Ser, i.e. transient potentiometry based D-Ser sensor using engineered DAAO showing quasi-direct electron transfer (DET) property. The previously produced G52V DAAO variant possesses dye-mediated dehydrogenase activity using artificial synthetic electron acceptors, while its oxidase activity is negligible. The enzyme was immobilized on an electrode and was modified with amine-reactive phenazine ethosulfate: the time dependent OCP change monitoring, transient potentiometry, provided rapid and sensitive sensor signals. D-Ser monitoring was followed at short resolution time (<1 s), with high sensitivity and in the 20 μM–30 mM range. D-Ser monitoring was then carried out in the artificial cerebrospinal fluid.



  • Separation and determination of cysteine enantiomers in plasma after derivatization with 4-fluoro-7-nitrobenzofurazan

Sabrina Ferré, Víctor González-Ruiz, Joséphine Zangari, Sergey Girel, Jean-Claude Martinou, Roccaldo Sardella, Serge Rudaz
Journal of Pharmaceutical and Biomedical Analysis (2022) Volume 209, 114539

Quantification of cysteine enantiomers in rodent plasma has been achieved by using 4-fluoro-7-nitrobenzofurazan derivatization of the target analytes. The enantioseparation was achieved in less than 3 min using a (R,R)-Whelk-O 1 stationary phase and isocratic elution: the derivatives were detected using negative ESI-MS in SRM mode. The method shows a response in the 0-300 µM and 0-125 µM range for D-cysteine and L-cysteine, respectively. The method was then applied for determining D- and L-cysteine in mouse plasma after D-cysteine administration.



  • Reconstruction of hyper-thermostable ancestral L-amino acid oxidase to perform deracemization to D-amino acids

Chiharu Ishida, Ryo Miyata, Fumihito Hasebe, Azusa Miyata, Shigenori Kumazawa, Sohei Ito, Shogo Nakano
ChemCatChem (2021) Volume 13, Issue 24, 5228-5235.

An hyper-thermostable L-amino acid oxidase (HTAncLAAO) was designed through a combination of manual sequence data mining and ancestral sequence reconstruction. Recombinant HTAncLAAO, overexpressed in E. coli cells, recognizes seven L-AAs as substrates, exhibits extremely high thermal stability and long-term stability. Deracemization was performed at 40 °C with a small amount of enzyme: a total of 0.4 mg (2 U) of HTAncLAAO is enough to deracemize three D,L-AAs at a preparative scale with high enantiomeric excess (>99 %).



  The D-amino acids International Research Center “DAAIR“ has been established in Gerenzano (Varese, Italy) in 2019 with the aim to support and perform scientific research projects and activities on the field of D-amino acids. The Center, located inside the Fondazione Istituto Insubrico Ricerca per la Vita, is aimed to represent a pole of excellence at international level for dissemination and research involving the D-amino acids (Director Silvia Sacchi).   The guiding principle is support the research projects aimed to investigate the involvement of D-amino acids in main physiological processes, from bacteria to humans. The ultimate goal is to actively participate to the elucidation of the mechanisms by which the D-amino acids perform specific functions, and to identify their presence and concentration in different organisms and compartments, also with regards to well-established functional states, with particular emphasis to pathological states. Understand the involvement of D-amino acids in important diseases as a way to set up novel therapeutic strategies.   Contacts:;;   
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