Activities and Regulation of Enzymes of Carbohydrate Metabolism in Spruce ( Picea abies)

Activities o f the glycolytic enzymes were determined in seedlings, callus cultures and cell sus­ pension cultures o f spruce ( Picea abies) (L.) (Karst). The rate-limiting enzymes o f the pathway were the hexokinases, ATP: phosphofructokinase, fructose-1,6-bisphosphatase and pyruvate kinase. Two phosphofructokinases were found: A T P : fructose-6-phosphate 1-phosphotransferase (PFK) and pyrophosphate :fructose6-phosphate 1-phosphotransferase (PFP). In the presence o f its activator fructose-2,6-bisphosphate, PFP had a 4 5-fold higher specific activity than PFK. PFP could be activated about 20-fold by fructose-2,6-bisphosphate at saturating concentrations o f the substrates (fructose6-phosphate and pyrophosphate). The increase o f Fmax was accompanied by a strong increase in the apparent affinity o f the enzyme for the substrates. Km for fructose-6-phosphate and pyrophosphate was 0.44 mM and 24 fiM, respectively. Ka for fructose-2,6-bisphosphate was 24 nM. In seedlings, specific activity o f the glycolytic enzymes was 3 0 -3 0 0 percent higher in the hypocotyls, except for fructose-1,6-bisphosphate aldolase, glyceraldehyde-3-phosphate de­ hydrogenase and phosphoglycerate kinase, their activity being 100150 percent higher in the cotyledons, This distribution remained unchanged during periods o f 2 16 weeks o f cultiva­ tion o f the seedlings. In callus cultures and in cell suspension cultures, grown mixotrophically with different car­ bohydrates, all enzymes were between 1and 7-fold higher than in autotrophically grown seed­ lings. Incubation o f seedlings in mineral salt mixture containing a carbohydrate resulted in a rapid coordinate increase o f the activities to the levels o f callusor cell suspension cultures. This induction required a carbohydrate and oxygen. During prolonged cultivation o f cell sus­ pension cultures, when carbohydrate became limiting, activity o f the enzymes slowly declined.