Evidence for phosphorylation/dephosphorylation control of phosphofructokinase from organs of the Anoxia‐Tolerant sea mussel Mytilus edulis

The transition from aerobic to anoxic conditions induced stable modifications of the kinetic and regulatory properties of 6‐phosphofructo‐1‐kinase (PFK) from the posterior adductor muscle and hepatopancreas of the marine bivalve mollusc Mytilus edulis L. Anoxia resulted in a reduction in the maximal activity of PFK, a decrease in affinity for fructose‐6‐P (S0.5 increased by 60% in muscle and twofold in hepatopancreas), reduced effects of activators (AMP, fructose‐2,6‐P2), and reduced inhibition by ATP (I50 increased 2–2.4 fold). Organ‐specific differences in PFK properties were greatest for enzyme activators, with the hepatopancreas enzyme showing much lower Ka values for AMP and fructose‐2,6‐P2 as well as activation by inorganic phosphate (Ka 0.07 mM aerobic, 0.11 mM anoxic). By contrast, the muscle enzyme was not affected by Pi and neither enzyme was inhibited by citrate. To determine if the anoxia‐induced modifications of PFK were due to covalent modification of the enzyme protein, PFK preparations were incubated with ATP + Mg2+ + protein kinase second messengers (cAMP, cGMP, or Ca2+ plus phorbol 12‐myristate 13‐acetate) or with Mg2+ + alkaline phosphatase. Protein kinase action on posterior adductor muscle PFK increased enzyme Vmax, and affinity for fructose‐6‐P. cAMP‐ or cGMP‐stimulated protein kinase had the same action on hepatopancreas PFK; but incubation with stimulators of protein kinase C had the opposite effect on the enzyme. Alkaline phosphatase treatment reversed the effects of in vitro phosphorylation. The data suggest that the glycolytic rate depression (reverse Pasteur effect) that occurs with the transition to anoxia in these facultative anaerobes is facilitated by reversible phosphorylation control over the activity of PFK.