A C-Terminal Phosphatase Module Conserved in Vertebrate CMP-Sialic Acid Synthetases Provides a Tetramerization Interface for the Physiologically Active Enzyme

• Published in: Journal of molecular biology Vol. 393; no. 1; pp. 83 - 97
• Main Authors: Oschlies, Melanie, Dickmanns, Achim, Haselhorst, Thomas, Schaper, Wiebke, Stummeyer, Katharina, Tiralongo, Joe, Weinhold, Birgit, Gerardy-Schahn, Rita, von Itzstein, Mark, Ficner, Ralf, Münster-Kühnel, Anja-K.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 16.10.2009
• Subjects: Amino Acid Sequence; Animals; CMP-sialic acid; CMP-sialic acid synthetase; Crystallography, X-Ray; HAD phosphatase; Kinetics; Mice; Models, Molecular; Molecular Sequence Data; N-Acylneuraminate Cytidylyltransferase - chemistry; N-Acylneuraminate Cytidylyltransferase - metabolism; Protein Multimerization; Protein Structure, Quaternary; Protein Structure, Tertiary; Sequence Alignment; sialic acid; α/β-type hydrolase fold; ManNAc-6P; CSS; STD; Neu5Ac; NT; CMP; CMP-sialic acid synthetase; HAD phosphatase; HAD; pNPP; SeMet; Sia; CIP; mCSS-NT; NPS; SEC; CT; MAD; PC; KDN; sialic acid; KDO; α/β-type hydrolase fold; CMP-sialic acid
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2009.08.003

The biosynthesis of sialic acid-containing glycoconjugates is crucial for the development of vertebrate life. Cytidine monophosphate-sialic acid synthetase (CSS) catalyzes the metabolic activation of sialic acids. In vertebrates, the enzyme is chimeric, with the N-terminal domain harboring the synthetase activity. The function of the highly conserved C-terminal domain (CSS-CT) is unknown. To shed light on its biological function, we solved the X-ray structure of murine CSS-CT to 1.9 Å resolution. CSS-CT is a stable shamrock-like tetramer that superimposes well with phosphatases of the haloacid dehalogenase superfamily. However, a region found exclusively in vertebrate CSS-CT appears to block the active-site entrance. Accordingly, no phosphatase activity was observed in vitro, which points toward a nonenzymatic function of CSS-CT. A computational three-dimensional model of full-length CSS, in combination with in vitro oligomerization studies, provides evidence that CSS-CT serves as a platform for the quaternary organization governing the kinetic properties of the physiologically active enzyme as demonstrated in kinetic studies.