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Serine racemase is responsible for the synthesis of D-serine, an endogenous co-agonist for N-methyl-D-aspartate receptor-type glutamate receptors (NMDARs). This pyridoxal 5-phosphatedependent enzyme is involved both in the reversible conversion of L-to D-serine and serine catabolism by ␣,␤-elimination of water, thereby regulating D-serine levels. Because D-serine affects NMDAR signaling throughout the brain, serine racemase is a promising target for the treatment of disorders related to NMDAR dysfunction. To provide a molecular basis for rational drug design the x-ray crystal structures of human and rat serine racemase were determined at 1.5-and 2.1-Å resolution, respectively, and in the presence and absence of the orthosteric inhibitor malonate. The structures revealed a fold typical of ␤-family pyridoxal 5-phosphate enzymes, with both a large domain and a flexible small domain associated into a symmetric dimer, and indicated a ligand-induced rearrangement of the small domain that organizes the active site for specific turnover of the substrate.N-Methyl-D-aspartate receptor-type glutamate receptors (NMDARs) 2 are a key component in glutamatergic transmission implicated in the development, function, and plasticity of the nervous system. In addition to the neurotransmitter glutamate, the activation of NMDARs requires the binding of either of the two endogenous co-agonists, glycine or D-serine, to the NMDAR "glycine modulatory site" (1, 2). Although both amino acids have similar potency as co-agonists, they display regional differences in modulating NMDAR function (3, 4). According to its distribution within the CNS, D-serine exerts its function predominantly in corticolimbic brain structures (5). This pivotal role of D-serine is supported by several experimental findings, including depletion studies in neuronal tissue preparations, indicating a strong reduction in NMDAR-mediated transmission and impaired synaptic plasticity in the absence of D-serine (6, 7).D-serine is produced by enzymatic conversion of L-to D-serine mediated by the pyridoxal 5Ј-phosphate (PLP)-containing enzyme serine racemase (SR). Mammalian SR was first purified from rat brain and functionally characterized in 1999 by Wolosker and colleagues (8). The enzyme is expressed in glial cells and neurons and constitutes the sole endogenous source for D-serine in mammals (9). Both human and rodent SR have been purified and extensively studied to generate comprehensive knowledge of the enzymatic characteristics and allosteric modulation by a number of agents such as ATP and Mg 2ϩ ions. In addition to serine isomerization, the enzyme catalyzes the ␣,␤-elimination of water from L-and D-serine to produce pyruvate and ammonia, thus, offering the possibility for SR not only to elevate but also to reduce the level of D-serine (5), a role that previously has mainly been attributed to D-amino acid oxidase (10).Recent data from SR-deficient mice strongly implicate this enzyme in the process of NMDAR-dependent plasticity and neurotoxicity. Knock-out mice were ...

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Transhydrogenase couples the redox (hydride-transfer) reaction between NAD(H) and NADP(H) to proton translocation across a membrane. The redox reaction is catalyzed at the interface between two components (dI and dIII) which protrude from the membrane. A complex formed from recombinant dI and dIII (the dI(2)dIII(1) complex) from Rhodospirillum rubrum transhydrogenase catalyzes fast single-turnover hydride transfer between bound nucleotides. In this report we describe three new crystal structures of the dI(2)dIII(1) complex in different nucleotide-bound forms. The structures reveal an asymmetry in nucleotide binding that complements results from solution studies and supports the notion that intact transhydrogenase functions by an alternating site mechanism. In one structure, the redox site is occupied by NADH (on dI) and NADPH (on dIII). The dihydronicotinamide rings take up positions which may approximate to the ground state for hydride transfer: the redox-active C4(N) atoms are separated by only 3.6 A, and the perceived reaction stereochemistry matches that observed experimentally. The NADH conformation is different in the two dI polypeptides of this form of the dI(2)dIII(1) complex. Comparisons between a number of X-ray structures show that a conformational change in the NADH is driven by relative movement of the two domains which comprise dI. It is suggested that an equivalent conformational change in the intact enzyme is important in gating the hydride-transfer reaction. The observed nucleotide conformational change in the dI(2)dIII(1) complex is accompanied by rearrangements in the orientation of local amino acid side chains which may be responsible for sealing the site from the solvent and polarizing hydride transfer.

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