Purification and regulatory properties of fructose 1,6-diphosphatase from Hydrogenomonas eutropha.
Fructose diphosphatase of Hydrogenomonas eutropha H 16, produced during autotrophic growth, was purified 247-fold from extracts of cells. The molecular weight of the enzyme was estimated to be 170,000. The enzyme showed a pH optimum of 8.5 in both crude extracts and purified preparation. The shape of the pH curve was not changed in the presence of ethylenediaminetetraacetic acid. The enzyme required Mg(2+) for activity. The MgCl(2) saturation curve was sigmoidal and the degree of positive cooperativity increased at lower fructose diphosphate concentrations. Mn(2+) can replace Mg(2+), but maximal activity was lower than that observed with Mg(2+) and the optimal concentration range was narrow. The fructose diphosphate curve was also sigmoidal. The purified enzyme also hydrolyzed sedoheptulose diphosphate but at a much lower rate than fructose diphosphate. The enzyme was not inhibited by adenosine 5'-monophosphate but was inhibited by ribulose 5-phosphate and adenosine 5'-triphosphate. Adenosine 5'-triphosphate did not affect the degree of cooperativity among the sites for fructose diphosphate. The inhibition by adenosine 5'-triphosphate was mixed and by ribulose 5-phosphate was noncompetitive. An attempt was made to correlate the properties of fructose diphosphatase from H. eutropha with its physiological role during autotrophic growth.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000645:Ammonium Sulfate; D002458:Cell Fractionation; D002474:Cell-Free System; D011232:Chemical Precipitation; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004492:Edetic Acid / Q000494:pharmacology; D006597:Fructose-Bisphosphatase / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D005636:Fructosephosphates / Q000096:biosynthesis; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D011479:Protamines; D011549:Pseudomonas / Q000201:enzymology*; D013403:Sugar Phosphates / Q000494:pharmacology
D005636:Fructosephosphates; D011479:Protamines; D013403:Sugar Phosphates; D008345:Manganese; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D006597:Fructose-Bisphosphatase; D008274:Magnesium; D000645:Ammonium Sulfate