MOLECULAR, KINETIC, AND IMMUNOLOGICAL PROPERTIES OF THE 6-PHOSPHOFRUCTOKINASE FROM THE GREEN-ALGA SELENASTRUM-MINUTUM - ACTIVATION DURING BIOSYNTHETIC CARBON FLOW

The ATP:D-fructose-6-phosphate 1-phosphotransferase (PFK) from Selenastrum minutum was purified to homogeneity. The purified plastid enzyme had a specific activity of 180 micromoles per milligram of protein per minute. It is a homomer with a subunit molecular weight of 70,000. The smallest enzymatically active form of the protein is a homotetramer of 280,000 daltons. The enzyme can, however, aggregate into different active forms, the largest of which has a molecular weight of more than 6 × 106. The pH optimum, regardless of aggregation state, is 7.25. The enzyme exhibits sigmoidal kinetics with respect to fructose-6-phosphate and hyperbolic kinetics with respect to ATP. Phosphate changes the sigmoidal fructose-6-phosphate saturation kinetics to hyperbolic. Phosphoenolpyruvate, 3-phosphoglycerate, 2-oxoglutarate, malate, citrate and ATP all inhibit the enzyme. The ratios of phosphoenolpyruvate and/or 3-PGA to phosphate are probably the most important factors regulating PFK activity in vivo. The enzyme cross-reacts with several antisera against both cytosolic and plastidic PFKs as well as against native potato pyrophosphate dependent phosphofructokinase suggesting that the algal PFK represents an evolutionarily primitive form.