Purification and properties of 3-hexulose phosphate synthase and phospho-3-hexuloisomerase from Methylococcus capsulatus.
3-Hexulose phosphate synthase and phospho-3-hexuloisomerase were purified 40- and 150-fold respectively from methane-grown Methylococcus capsulatus. The molecular weights of the enzymes were approximately 310000 and 67000 respectively, as determined by gel filtration. Dissociation of 3-hexulose phosphate synthase into subunits of molecular weight approx. 49000 under conditions of low pH or low ionic strength was observed. Within the range of compounds tested, 3-hexulose phosphate synthase is specific for formaldehyde and d-ribulose 5-phosphate (forward reaction) and d-arabino-3-hexulose 6-phosphate (reverse reaction), and phospho-3-hexuloisomerase is specific for d-arabino-3-hexulose 6-phosphate (forward reaction) and d-fructose 6-phosphate (reverse reaction). A bivalent cation is essential for activity and stability of 3-hexulose phosphate synthase; phospho-3-hexuloisomerase is inhibited by many bivalent cations. The pH optima of the two enzymes are 7.0 and 8.3 respectively and the equilibrium constants are 4.0x10(-5)m and 1.9x10(2)m respectively. The apparent Michaelis constants for 3-hexulose phosphate synthase are: d-ribulose 5-phosphate, 8.3x10(-5)m; formaldehyde, 4.9x10(-4)m; d-arabino-3-hexulose 6-phosphate, 7.5x10(-5)m. The apparent Michaelis constants for phospho-3-hexuloisomerase are: d-arabino-3-hexulose 6-phosphate, 1.0x10(-4)m; d-fructose 6-phosphate, 1.1x10(-3)m.
D000446:Aldehyde-Lyases / Q000302:isolation & purification*; D019747:Aldose-Ketose Isomerases; D002238:Carbohydrate Epimerases / Q000302:isolation & purification*; D003035:Cobalt; D006863:Hydrogen-Ion Concentration; D007661:Ketoses; D007700:Kinetics; D008274:Magnesium; D008345:Manganese; D008772:Methylococcaceae / Q000201:enzymology*; D008970:Molecular Weight; D013403:Sugar Phosphates; D015032:Zinc
D007661:Ketoses; D013403:Sugar Phosphates; D003035:Cobalt; D008345:Manganese; D000446:Aldehyde-Lyases; C021355:hexose phosphate synthetase; D002238:Carbohydrate Epimerases; D019747:Aldose-Ketose Isomerases; C023918:phospho-3-hexuloisomerase; D008274:Magnesium; D015032:Zinc