Fructose-2,6-bisphosphate metabolism in permeabilized yeast cells

Several methods for the permeabilization of Saccharomyces cerevisiae M1 were compared. Cells were permeabilized in the presence of 3% toluene/mercaptoethanol, and the activities of 6-phosphofructo-2-kinase, fructose-2,6-bisphosphatase and alkaline phosphatase were measured during growth of yeast on glucose. In the exponential phase of growth, the specific activities of 6-phosphofructo-2-kinase and fructose-2,6-bisphosphatase decrease significantly. The specific activities of 6-phosphofructo-2-kinase and high-affinity fructose-2,6-bisphosphatase increase again during the transition phase and reach maximum values in the stationary phase. In contrast to the specific activities, the activity concentrations of 6-phosphofructo-2-kinase and fructose-2,6-bisphosphatase remain nearly constant in the exponential phase, but increase in the transition and the stationary growth phase. The concentration of fructose-2,6-bisphosphate drops from about 6 microM in the exponential phase to very low levels in the transition phase, but increases slightly in the stationary phase. In Saccharomyces cerevisiae M1 several fructose-2,6-bisphosphate degrading activities were measured differing in the behaviour during growth on glucose, in the pH-optimum and the inhibition by fructose-6-phosphate.