Species: Rattus norvegicus
Subunit composition of N-acetylneuraminate 9-phosphate synthase = [N-acetylneuraminate 9-phosphate synthase subunit]2
The native apparent molecular mass was determined by gel filtration chromatography [Chen02a].
This enzyme catalyzes a critical reaction in sialic acid biosynthesis, the condensation of phosphoenolpyruvate with N-acetyl-D-mannosamine 6-phosphate. Unlike the bacterial enzyme that uses N-acetyl-β-D-mannosamine as substrate, the animal enzyme uses the 6-phosphorylated derivative to produce N-acetyl-β-neuraminate 9-phosphate (see pathways CMP-N-acetylneuraminate biosynthesis I (eukaryotes) and CMP-N-acetylneuraminate biosynthesis II (bacteria)).
A key difference also exists in the human sialic acid synthase versus rat (this enzyme) and mouse N-acetylneuraminate 9-phosphate synthase enzymes. The human enzyme can also utilize α-D-mannose 6-phosphate to produce 2-keto-3-deoxy-D-glycero-D-galacto-nononate 9-phosphate in this reaction (see pathway CMP-2-keto-3-deoxy-D-glycero-D-galacto-nononate biosynthesis), whereas the rodent enzymes cannot (reviewed in [Tanner05]). Mutagenesis studies of the recombinant human enzyme revealed that a single mutation resulting in substitution of Thr for Met at position 42 eliminated the 2-keto-3-deoxy-D-glycero-D-galacto-nononate 9-phosphate synthase activity without affecting N-acetyl-β-neuraminate 9-phosphate synthase activity [Hao06].
The native rat liver enzyme was purified to homogeneity and was determined to be a homodimer
[Chen02a]. Although the human
sialic acid synthase
[Lawrence00] and mouse
N-acetylneuraminate 9-phosphate synthase
[Nakata00] enzymes have been cloned, the rat enzyme has not.
The apparent molecular mass of the subunit was determined by SDS-PAGE [Chen02a].
Molecular Weight of Polypeptide: 37.0 kD (experimental) [Chen02a]
Molecular Weight of Multimer: 75.0 kD (experimental) [Chen02a]
Enzymatic reaction of: N-acetylneuraminate 9-phosphate synthase
Synonyms: N-acetylneuraminate phosphate synthase, N-acetylneuraminic acid phosphate synthase, N-acetylneuraminic acid 9-phosphate synthase, sialic acid (N-acetylneuraminic acid) synthase, sialic acid synthase
EC Number: 18.104.22.168phosphoenolpyruvate + N-acetyl-D-mannosamine 6-phosphate + H2O → N-acetyl-β-neuraminate 9-phosphate + phosphate
The direction shown, i.e. which substrates are on the left and right sides, is in accordance with the Enzyme Commission system.
The reaction is favored in the direction shown.
The enzyme was highly specific for N-acetyl-D-mannosamine 6-phosphate and did not utilize D-glucosamine 6-phosphate, β-D-fructofuranose 6-phosphate, N-acetyl-α-D-glucosamine 1-phosphate, N-acetyl-D-glucosamine 6-phosphate, or N-acetyl-β-D-mannosamine as substrate. pyruvate could not replace phosphoenolpyruvate [Chen02a].
The rat enzyme also did not use α-D-mannose 6-phosphate as substrate and therefore lacked 2-keto-3-deoxy-D-glycero-D-galacto-nononate 9-phosphate synthase activity [Chen02a].
The mechanism of this condensation reaction remains to be completely elucidated, although possible mechanisms have been considered that involve either C-O, or P-O bond cleavage (reviewed in [Tanner05]).N-ethylmaleimide [Chen02a], 5,5'-dithio-bis-2-nitrobenzoate [Chen02a]
Chen02a: Chen H, Blume A, Zimmermann-Kordmann M, Reutter W, Hinderlich S (2002). "Purification and characterization of N-acetylneuraminic acid-9-phosphate synthase from rat liver." Glycobiology 12(2);65-71. PMID: 11886839
Hao06: Hao J, Vann WF, Hinderlich S, Sundaramoorthy M (2006). "Elimination of 2-keto-3-deoxy-D-glycero-D-galacto-nonulosonic acid 9-phosphate synthase activity from human N-acetylneuraminic acid 9-phosphate synthase by a single mutation." Biochem J 397(1);195-201. PMID: 16503877
Lawrence00: Lawrence SM, Huddleston KA, Pitts LR, Nguyen N, Lee YC, Vann WF, Coleman TA, Betenbaugh MJ (2000). "Cloning and expression of the human N-acetylneuraminic acid phosphate synthase gene with 2-keto-3-deoxy-D-glycero- D-galacto-nononic acid biosynthetic ability." J Biol Chem 275(23);17869-77. PMID: 10749855
Nakata00: Nakata D, Close BE, Colley KJ, Matsuda T, Kitajima K (2000). "Molecular cloning and expression of the mouse N-acetylneuraminic acid 9-phosphate synthase which does not have deaminoneuraminic acid (KDN) 9-phosphate synthase activity." Biochem Biophys Res Commun 273(2);642-8. PMID: 10873658
©2016 SRI International, 333 Ravenswood Avenue, Menlo Park, CA 94025-3493