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.

Magnesium and calcium ions differentially affect human serine racemase activity and modulate its quaternary equilibrium toward a tetrameric form / Bruno, Stefano; Margiotta, Marilena; Marchesani, Francesco; Paredi, Gianluca; Orlandi, Valentina; Faggiano, Serena; Ronda, Luca; Campanini, Barbara; Mozzarelli, Andrea. - In: BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS. - ISSN 1570-9639. - 1865:4(2017), pp. 381-387. [10.1016/j.bbapap.2017.01.001]

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

BRUNO, Stefano
;
MARGIOTTA, MARILENA;MARCHESANI, FRANCESCO;PAREDI, Gianluca;FAGGIANO, Serena;RONDA, Luca;CAMPANINI, Barbara;MOZZARELLI, Andrea
2017

Abstract

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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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11381/2823648
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