Isolation and characterization of a pyrophosphate-dependent phosphofructokinase from Propionibacterium shermanii.
A pyrophosphate-dependent phosphofructokinase (pyrophosphate; D-fructose-6-phosphate-1-phosphotransferase) has been purified and characterized from extracts of Propionibacterium shermanii. The enzyme catalyzes the transfer of phosphate from pyrophosphate to fructose 6-phosphate to yield fructose-1,6-P2 and phosphate. This unique enzymatic activity was observed initially in Entamoeba histolytica (Reeves, R.E., South, D.J., Blytt, H.G., and Warren, L. G. (1974) J. Biol. Chem. 249, 7734-7741). This is the third pyrophosphate-utilizing enzyme that these two diverse organisms have in common. The others are phosphoenolpyruvate carboxytransphosphorylase and pyruvate phosphate dikinase. The PPi-phosphofructokinase from P. shermanii is specific for fructose-6-P and fructose-1,6-P2, no other phosphorylated sugars were utilized. Phosphate could be replaced by arsenate. The Km values are: phosphate, 6.0 X 10(-4) M; fructose-1, 6-P2, 5.1 X 10(-5) M; pyrophosphate, 6.9 X 10(-5) M; and fructose-6-P, 1.0 X 10(-4) M. The S20w is 5.1 S. The molecular weight of the native enzyme is 95,000. Sodium dodecyl sulfate electrophoresis of the enzyme showed a single band migrating with an Rf corresponding to a molecular weight of 48,000. Extracts of P. shermanii have PPi-phosphofructokinase activity approximately 6 times greater than ATP-phosphofructokinase and 15 to 20 times greater than fructose diphosphatase activities. It is proposed that (a) PPi may replace ATP in the formation of fructose-1-6-P2 when the organism is grown on glucose and (b) when the organism is grown on lactate or glycerol the conversion of fructose-1,6-P2 to fructose-6-P during gluconeogenesis may occur by phosphorolysis rather than hydrolysis.
D011756:Diphosphates / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D010732:Phosphofructokinase-1 / Q000302:isolation & purification / Q000378:metabolism*; D011424:Propionibacterium / Q000187:drug effects / Q000201:enzymology*; D013329:Structure-Activity Relationship
D011756:Diphosphates; D008345:Manganese; D010732:Phosphofructokinase-1; D008274:Magnesium