Resolution and characterization of multiple cytosolic phosphatases capable of hydrolyzing fructose‐1,6‐bisphosphate in spinach and soybean leaves

The apparent activity of cytoplasmic fructose bisphosphatase (EC 3.1.3.11) in crude extracts of spinach (Spinacia oleracea L.) and soybean (Glycine max [L.] Merr.) leaves was only partially dependent on Mg²⁺. At least two major non‐chloroplastic fructose bisphosphatases that differed in dependence on Mg²⁺ were chromatographically resolved from spinach leaves. The Mg²⁺‐dependent enzyme had an apparent Michaelis constant of 4 μM for fructose‐1,6‐P₂, was highly specific, and was strongly inhibited by fructose‐2,6‐P₂. Enzyme activity was inhibited by physiological levels of fructose‐6‐P. Both species also contained at least one major enzyme, the activity of which was independent of Mg²⁺. These enzymes had pH optima near neutrality, Michaelis constants of 25 to 30 μM for fructose‐1,6‐P₂, and were inhibited by AMP. Although hexose monophosphates were not metabolized, the enzymes were not specific for fructose‐1,6‐P₂: phosphate was released from phosphoenolpyruvate and ribulose‐1, 5‐P₂, and with fructose‐1,6‐P₂, as substrate, Pi release was about 1.5‐fold greater than fructose‐6‐P production. It is concluded that only the Mg²⁺‐dependent fructose bisphosphatase, previously characterized, functions in the photosynthetic sucrose formation pathway. Inhibition of the Mg²⁺‐dependent enzyme by fructose‐6‐P may be involved in regulation of sucrose formation.