Serine racemase is a pyridoxal 5'‑phosphate dependent enzyme responsible for the synthesis of d‑serine, a neuromodulator of the NMDA receptors. Its activity is modulated by several ligands, including ATP, divalent cations and protein interactors. The murine orthologue is inhibited by S-nitrosylation at Cys113, a residue adjacent to the ATP binding site. We found that the time course of inhibition of human serine racemase by S-nitrosylation is markedly biphasic, with a fast phase associated with the reaction of Cys113. Unlike the murine enzyme, two additional cysteine residues, Cys269, unique to the human orthologue, and Cys128 were also recognized as S-nitrosylation sites through mass spectrometry and site-directed mutagenesis. The effect of S-nitrosylation on the fluorescence of tryptophan residues and on that of the pyridoxal phosphate cofactor indicated that S-nitrosylation produces a partial interruption of the cross-talk between the ATP binding site and the active site. Overall, it appears that the inhibition results from a conformational change rather than the direct displacement of ATP.

Human serine racemase is nitrosylated at multiple sites / Marchesani, Francesco; Bruno, Stefano; Paredi, Gianluca; Raboni, Samanta; Campanini, Barbara; Mozzarelli, Andrea. - In: BIOCHIMICA ET BIOPHYSICA ACTA. - ISSN 0006-3002. - 1866:7(2018), pp. 813-821. [10.1016/j.bbapap.2018.01.009]

Human serine racemase is nitrosylated at multiple sites

Marchesani, Francesco;Bruno, Stefano
;
Raboni, Samanta;Campanini, Barbara;Mozzarelli, Andrea
2018

Abstract

Serine racemase is a pyridoxal 5'‑phosphate dependent enzyme responsible for the synthesis of d‑serine, a neuromodulator of the NMDA receptors. Its activity is modulated by several ligands, including ATP, divalent cations and protein interactors. The murine orthologue is inhibited by S-nitrosylation at Cys113, a residue adjacent to the ATP binding site. We found that the time course of inhibition of human serine racemase by S-nitrosylation is markedly biphasic, with a fast phase associated with the reaction of Cys113. Unlike the murine enzyme, two additional cysteine residues, Cys269, unique to the human orthologue, and Cys128 were also recognized as S-nitrosylation sites through mass spectrometry and site-directed mutagenesis. The effect of S-nitrosylation on the fluorescence of tryptophan residues and on that of the pyridoxal phosphate cofactor indicated that S-nitrosylation produces a partial interruption of the cross-talk between the ATP binding site and the active site. Overall, it appears that the inhibition results from a conformational change rather than the direct displacement of ATP.
Human serine racemase is nitrosylated at multiple sites / Marchesani, Francesco; Bruno, Stefano; Paredi, Gianluca; Raboni, Samanta; Campanini, Barbara; Mozzarelli, Andrea. - In: BIOCHIMICA ET BIOPHYSICA ACTA. - ISSN 0006-3002. - 1866:7(2018), pp. 813-821. [10.1016/j.bbapap.2018.01.009]
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11381/2841533
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