Phosphofructokinase (ATP:D-fructose-6phos-phate 1-phosphotransferase, EC 2.7.1.11) was partially purified the livers of genetically diabetic mice (C57BL/Ksj-db) and their lean littermates (C57BL/KsJ). These genetically diabetic mice have been shown to be hyperglucagonemic and to exhibit symptoms resembling those of maturity-onset diabetes in humans. Two isozymes of phosphofructokinase were obtained after DEAE-Sephadex chromatography of extracts of livers from either normal or diabetic animals. One of these isozymes, peak II, from the genetically diabetic mice was shown to be more sensitive to ATP inhibition at physiological pH than the peak II isozyme from the normal animals. In addition, the peak II isozyme from the diabetic mice exhibited decreased affinity for fructose 6-phosphate. The (4). Enhanced gluconeogenesis appears to be a major factor in the hyperglycemia of diabetic mice. Studies of gluconeogenic fluxes and metabolite levels in diabetic mice indicate that the increased liver gluconeogenesis is likely to be due to a facilitation of the fructose 1,6-bisphosphatase (D-fructose-1,6-bisphosphate 1-phosphohydrolase, EC 3.1.3.11) reaction or a restraint of the phosphofructokinase (ATP:D-fructose-6-phosphate 1-phosphotransferase, EC 2.7.1.11) reaction or both (5). Studies on gluconeogenic rates in isolated rat liver cells also led to the proposal that glucagon enhances gluconeogenesis and inhibits glycolysis at the site of fructose 1,6-bisphosphatase and phosphofructokinase (6).With this background it appeared to us that a comparative study of both phosphofructokinase and fructose 1,6-bisphosphatase from the livers of C57BL/KsJ normal and diabetic hyperglucagonemic mice could provide an insight into the mode of action of glucagon at the metabolic steps catalyzed by the above enzymes.The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.
2497A previous study of fructose 1,6-bisphosphatase isolated from the livers of diabetic mice and from the normal controls showed no difference in either molecular or enzymatic properties between the two purified enzymes (7). In this communication we show that phosphofructokinase isolated from the livers of diabetic mice is more sensitive to ATP inhibition than is the enzyme isolated from the livers of normal controls and therefore may represent a physiologically less active form of phosphofructokinase. These results are entirely in accord with three recent independent reports which have indicated decreased liver phosphofructokinase activity after either administration of glucagon to rats (8) or addition of glucagon to isolated rat hepatocytes (9, 10).
MATERIALS AND METHODSFructose 6-phosphate, fructose 1,6-bisphosphate, ATP, NADH, DEAE-Sephadex A-25, and aldolase were purchased from Sigma. Triose-phosphate isomerase and a-glycerophosphate dehydrogenase were from Boehringer Mannheim. Ammonium sulfate was from ...
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