Magnesium and calcium ions differentially affect human serine racemase activity and modulate its quaternary equilibrium toward a tetrameric form

• Published in: Biochimica et biophysica acta. Proteins and proteomics Vol. 1865; no. 4; pp. 381 - 387
• Main Authors: Bruno, Stefano, Margiotta, Marilena, Marchesani, Francesco, Paredi, Gianluca, Orlandi, Valentina, Faggiano, Serena, Ronda, Luca, Campanini, Barbara, Mozzarelli, Andrea
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-04-2017
• Subjects: Adenosine Triphosphate - chemistry; Allosteric regulation; Binding Sites; Calcium - chemistry; Enzyme regulation; Humans; Magnesium - chemistry; Metal-protein interaction; Metalloproteins; Protein Structure, Quaternary; Racemases and Epimerases - chemistry; Serine racemase; TEA; LDH; AMP; Allosteric regulation; Metal-protein interaction; hSR; Enzyme regulation; AMw; ATP; ADP; Serine racemase; Metalloproteins
• ISSN: 1570-9639; 1878-1454
• DOI: 10.1016/j.bbapap.2017.01.001

Serine racemase is the pyridoxal 5′-phosphate dependent enzyme that catalyzes both production and catabolism of d-serine, a co-agonist of the NMDA glutamate receptors. Mg2+, or, alternatively, Ca2+, activate human serine racemase by binding both at a specific site and – as ATP-metal complexes – at a distinct ATP binding site. We show that Mg2+ and Ca2+ bind at the metal binding site with a 4.5-fold difference in affinity, producing a similar thermal stabilization and partially shifting the dimer-tetramer equilibrium in favour of the latter. The ATP-Ca2+ complex produces a 2-fold lower maximal activation in comparison to the ATP-Mg2+ complex and exhibits a 3-fold higher EC50. The co-presence of ATP and metals further stabilizes the tetramer. In consideration of the cellular concentrations of Mg2+ and Ca2+, even taking into account the fluctuations of the latter, these results point to Mg2+ as the sole physiologically relevant ligand both at the metal binding site and at the ATP binding site. The stabilization of the tetramer by both metals and ATP-metal complexes suggests a quaternary activation mechanism mediated by 5′-phosphonucleotides similar to that observed in the distantly related prokaryotic threonine deaminases. This allosteric mechanism has never been observed before in mammalian fold type II pyridoxal 5′-phosphate dependent enzymes.
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•Mg2+ activates allosterically serine racemase in competition with Ca2+.•The ATP-Ca2+ complex produces half the activation brought about by the ATP-Mg2+ complex.•Ca2+ and Mg2+ stabilize a tetrameric form of the enzyme.•ATP, in complex with Ca2+ or Mg2+, further shifts the quaternary equilibrium toward the tetrameric form.•Ca2+ and Mg2+ increase the thermal stability of the enzyme.