Abstract
There is currently great interest in human serine racemase, the enzyme responsible for producing the NMDA co-agonist d-serine. Reported correlation of d-serine levels with disorders including Alzheimer's disease, ALS, and ischemic brain damage (elevated d-serine) and schizophrenia (reduced d-serine) has further piqued this interest. Reported here is a structure/activity relationship study of position Ser84, the putative re-face base. In the most extreme case of functional reprogramming, the S84D mutant displays a dramatic reversal of β-elimination substrate specificity in favor of l-serine over the normally preferred l-serine-O-sulfate (∼1200-fold change in kcat/Km ratios) and l (l-THA; ∼5000-fold change in kcat/Km ratios) alternative substrates. On the other hand, the S84T (which performs l-Ser racemization activity), S84A (good kcat but high Km for l-THA elimination), and S84N mutants (nearly WT efficiency for l-Ser elimination) displayed intermediate activity, all showing a preference for the anionic substrates, but generally attenuated compared with the native enzyme. Inhibition studies with l-erythro-β-hydroxyaspartate follow this trend, with both WT serine racemase and the S84N mutant being competitively inhibited, with Ki = 31 ± 1.5 μm and 1.5 ± 0.1 mm, respectively, and the S84D being inert to inhibition. Computational modeling pointed to a key role for residue Arg-135 in binding and properly positioning the l-THA and l-serine-O-sulfate substrates and the l-erythro-β-hydroxyaspartate inhibitor. Examination of available sequence data suggests that Arg-135 may have originated for l-THA-like β-elimination function in earlier evolutionary variants, and examination of available structural data suggests that a Ser84-H2O-Lys114 hydrogen-bonding network in human serine racemase lowers the pKa of the Ser84re-face base.
Highlights
The discovery of D-serine in the brain and its importance in modulating NMDA receptor activity provided the first bona fide example of a D-amino acid in human biology
L-Serine serves a central role in maintaining redox homeostasis, because all glutathione equivalents originate in the L-serine backbone, with the sulfur atom from dietary methionine being installed at the -carbon through the sequential action of two additional pyridoxal phosphate (PLP)-dependent enzymes, cystathionine -synthase (CBS) and cystathionine eliminase
It was found that improved human serine racemase (hSR) solubility could be achieved by expressing the protein as an N-terminal maltose-binding protein (MBP) fusion construct (MBP– hSR)
Summary
The discovery of D-serine in the brain and its importance in modulating NMDA receptor activity provided the first bona fide example of a D-amino acid in human biology. The observation that all D-serine apparently originates in L-serine added another significant branch to the complex metabolic network associated with L-serine and an important new signaling function for the amino acid (Fig. 1). From the point of view of neuronal signaling, both D-serine and glycine serve as co-agonists of the NMDA receptor (Fig. 2), binding at the “glycine site” but with the observation that D-serine is a more potent agonist than glycine itself, showing efficacy at several orders of magnitude lower concentration in a seminal study by Ascher and co-workers [2] in a rat hypoglossal motoneuron system [2,3,4]. Whereas it had previously been thought that D-serine is biosynthesized by SR in astroglial cells, more recent evidence indicates that L-serine produced in the astroglia from 3-phosphoglycerate is actively shuttled to the neurons where SR is present and converts the L-serine to D-serine [5, 6]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
