Effect of anoxia on the kinetic properties of pyruvate kinase and phosphofructokinase,and on glycogen phosphorylase activity in marine worms and earth worms

Summary The kinetic properties of PK and PFK were studied in aerobic versus 12-hours anoxic marine worms Hedistae(=Nereis) diversicolor and Diopatra neapolitana and earth worms Allolobophora calliginosa and Eisenia foetida. The total glycogen phosphorylase (a+b) activity and the percentage of active a form were also measured in the marine and earth worms under the same conditions. Anoxia exposure did not result in any significant changes of kinetic parameters of PK and total activities of glycogen phosphorylase from marine worms, but it altered the kinetic characteristics of PFK from H. diversicolor. Chromatographical studies showed that PK from both aerobic and anoxic marine worms is eluted from DEAE-cellulose as a single peak at 50 mM KCl. In contrast to marine worms, however, anoxia caused a marked change in kinetic properties of PK from both earth worms, resulting in a reduction of enzyme affinity for its substrate PEP. In addition, the enzyme existed in both earth worms in two distinct variants eluted from DEAE-cellulose column as peak I and peak II at 50 mM and 150 mM KCl, respectively. The ratio of enzyme units (peak I/peak II) was reduced significantly after 12 h of anoxia, indicating that these two peaks are interconvertible. Anoxia also caused a reduction of total glycogen phosphorylase activity in E. foetida and lowered the percentage of active a form of the enzyme by approximately 50% in both earth worms. Kinetic properties of PFK from both earth worms were not significantly affected by anoxia. However, their low Ka values for F-2,6-P₂ imply that this effector may play an important role in PFK control in earth worms under anoxia.Abbreviations F6P fructose-6-phosphate - FBP fructose-1,6-bisphosphate - F-2,6-P fructose-2,6-bisphosphate - PEP phosphoenoylpruvate - PFK 6-phosphofructo-1-kinase (E.C.2.7.1.11) - PK pyruvate kinase (E.C.2.7.1.40) - Pi inorganic phosphate - PMSF phenyl methylsulfonyl fluoride