N-methyl-d-aspartate receptors (NMDARs) are located in neuronal cell membranes at synaptic and extrasynaptic locations, where they are believed to mediate distinct physiological and pathological processes. Activation of NMDARs requires glutamate and a coagonist whose nature and impact on NMDAR physiology remain elusive. We report that synaptic and extrasynaptic NMDARs are gated by different endogenous coagonists, d-serine and glycine, respectively. The regionalized availability of the coagonists matches the preferential affinity of synaptic NMDARs for d-serine and extrasynaptic NMDARs for glycine. Furthermore, glycine and d-serine inhibit NMDAR surface trafficking in a subunit-dependent manner, which is likely to influence NMDARs subcellular location. Taking advantage of this coagonist segregation, we demonstrate that long-term potentiation and NMDA-induced neurotoxicity rely on synaptic NMDARs only. Conversely, long-term depression requires both synaptic and extrasynaptic receptors. Our observations provide key insights into the operating mode of NMDARs, emphasizing functional distinctions between synaptic and extrasynaptic NMDARs in brain physiology.

D-Amino acid oxidase (DAAO) is a FAD-containing flavoenzyme that catalyzes the oxidative deamination of D-isomers of neutral and polar amino acids. This enzymatic activity has been identified in most eukaryotic organisms, the only exception being plants. In the various organisms in which it does occur, DAAO fulfills distinct physiological functions: from a catabolic role in yeast cells, which allows them to grow on D-amino acids as carbon and energy sources, to a regulatory role in the human brain, where it controls the levels of the neuromodulator D-serine. Since 1935, DAAO has been the object of an astonishing number of investigations and has become a model for the dehydrogenase-oxidase class of flavoproteins. Structural and functional studies have suggested that specific physiological functions are implemented through the use of different structural elements that control access to the active site and substrate/product exchange. Current research is attempting to delineate the regulation of DAAO functions in the contest of complex biochemical and physiological networks.

Human genes coding for pLG72 and D-amino acid oxidase have recently been linked to the onset of schizophrenia. pLG72 was proposed as an activator of the human FAD-containing flavoprotein D-amino acid oxidase (hDAAO). In the brain this oxidizes D-serine, a potent activator of N-methyl-D-aspartate receptor. We have investigated the mechanistic regulation of hDAAO by pLG72. Immunohistochemical analyses revealed that hDAAO and pLG72 are both expressed in astrocytes of the human cortex, where they most likely interact, considering their partial overlapping subcellular distribution and their coimmunoprecipitation. We demonstrated that the specific in vitro interaction of the two proteins yields a complex composed of 2 hDAAO homodimers and 2 pLG72 molecules. Binding of pLG72 did not affect the kinetic properties and FAD binding ability of hDAAO; instead, a time-dependent loss of hDAAO activity in the presence of an excess of pLG72 was found. The binding affects the tertiary structure of hDAAO, altering the amount of the active form. We finally demonstrated that overexpression of hDAAO in glioblastoma cells decreases the levels of D-serine, an effect that is null when pLG72 is coexpressed. These data indicate that pLG72 acts as a negative effector of hDAAO. Therefore, a decrease in the synaptic concentration of D-serine as the result of an anomalous increase in hDAAO activity related to hypoexpression of pLG72 may represent a molecular mechanism by which hDAAO and pLG72 are involved in schizophrenia susceptibility.Schizophrenia is one of the most widely spread psychiatric disorders; it is a complex disease or, more likely, a group of related illnesses to which an individual has a strong genetic predisposition (1). Among the identified schizophrenia susceptibility genes (2), the gene G72 encodes for several splicing isoforms; pLG72 represents the longest open reading frame (153 amino acids), which is mainly expressed in brain (3). G72 is present only in primates: there are no homologues of this gene in databases nor has sequence analysis of the putative open reading frame revealed any likely function (2, 3). Yeast twohybrid experiments using pLG72 as bait identified D-amino acid oxidase (EC 1.4.3.3, DAAO 4 ) on 12q24 as a putative interacting partner, and preliminary functional measurements showed that pLG72 should function as an in vitro activator of pig kidney DAAO (pkDAAO) (3). DAAO is a FAD-containing flavoenzyme that catalyzes the oxidative deamination of D-amino acids to the corresponding ␣-keto acids, hydrogen peroxide and ammonia (4, 5).Based on current findings we can hypothesize that in brain, the physiological role of DAAO is to modulate the levels of D-serine, an important glial-derived messenger that acts as the endogenous allosteric modulator of the glutamatergic NMDA receptor subtype (6 -8). D-and L-serine can be reversibly isomerized in astrocytic glia, which unsheathes synapses, by serine racemase. Compelling evidence has indicated that glutamate neurotransmission hypofunction is associated with symptoms...

D D-Amino acid oxidase (DAAO) has been proposed to be involved in the oxidation of D D-serine, an allosteric activator of the NMDA-type glutamate receptor in the brain, and to be associated with the onset of schizophrenia. The recombinant human DAAO was expressed in Escherichia coli and was isolated as an active homodimeric flavoenzyme. It shows the properties of the dehydrogenase-oxidase class of flavoproteins, possesses a low kinetic efficiency, and follows a ternary complex (sequential) kinetic mechanism. In contrast to the other known DAAOs, the human enzyme is a stable homodimer even in the apoprotein form and weakly binds the cofactor in the free form.