Journal
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS
Volume 1865, Issue 4, Pages 381-387Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbapap.2017.01.001
Keywords
Serine racemase; Allosteric regulation; Metal-protein interaction; Enzyme regulation; Metalloproteins
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Funding
- University of Parma (FIL)
- French Infrastructure for Integrated Structural Biology (FRISBI) by Instruct as part of the European Strategy Forum on Research Infrastructures (ESFRI) [ANR-10-INSB-05-01]
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Serine racemase is the pyridoxal 5'-phosphate dependent enzyme that catalyzes both production and catabolism of o-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 Mgt} 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. (C) 2017 Elsevier B.V. All rights reserved.
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