Rabbit brain phosphofructokinase. Comparison of regulatory properties with those of other phosphofructokinase isozymes

M. Y. Tsai, R. G. Kemp

Research output: Contribution to journalArticle

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Abstract

A procedure was developed for the purification of phosphofructokinase from rabbit brain extracts. The enzyme was purified more than 100 fold with a yield of greater than 50%. It was found to have the same distribution of multiple molecular forms as found in the total brain extract, including one molecular species that is different from the more extensively studied muscle (heart) and liver (erythrocyte) isozymes. The regulatory properties of brain phosphofructokinase were compared with those of muscle and liver phosphofructokinases at pH 7.1 and 7.4. All 3 enzymes were inhibited less at pH 7.4 than at 7.1. At pH 7.1 the sensitivity to ATP as an inhibitor decreased in the order liver > muscle > brain. At pH 7.4, the relative sensitivities of brain and muscle phosphofructokinases were reversed with the brain enzyme being inhibited at slightly lower concentrations of ATP. Other inhibitors acted synergistically with ATP. Sensitivity to inhibition by 3 phosphoglycerate and phosphoenolpyruvate decreased in the order muscle > brain > liver. With 2,3 diphosphoglycerate as an inhibitor, the sensitivity decreased in the order liver > muscle > brain. Muscle phosphofructokinase was the most sensitive to citrate inhibition. Creatine phosphate, a potent inhibitor of muscle phosphofructokinase, was completely ineffective as an inhibitor of liver and brain phosphofructokinase. The actions of activators were evaluated at pH 7.4 in the presence of inhibitory concentrations of ATP. The 3 enzymes were almost equally responsive to the deinhibiting action of AMP and cyclic adenosine 3':5' monophosphate, although the enzyme from liver requires slightly higher concentrations. Muscle phosphofructokinase was the least sensitive to deinhibition by inorganic phosphate. The results are discussed with respect to the varying modes of carbohydrate metabolism in the different tissues taking into consideration the tissue concentrations of the effectors and the variations of concentration in different physiological states. The enzymes from brain and liver are apparently chiefly controlled by the relative amounts of the adenine nucleotides and inorganic phosphate. An exception may be the erythrocyte, which has the same isozyme that is present in liver. Here, 2,3 diphosphoglycerate may play an important role in regulation. Muscle phosphofructokinase also responds to varying levels of adenine nucleotides but in vivo it is most likely that the principal regulatory effectors are creatine phosphate, citrate, and possibly, 3 phosphoglycerate.

Original languageEnglish (US)
Pages (from-to)6590-6596
Number of pages7
JournalJournal of Biological Chemistry
Volume249
Issue number20
StatePublished - Dec 1 1974

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Phosphofructokinases
Isoenzymes
Muscle
Liver
Brain
Rabbits
Muscles
Enzymes
Adenosine Triphosphate
2,3-Diphosphoglycerate
Phosphocreatine
Adenine Nucleotides
Citric Acid
Erythrocytes
Phosphates
Tissue
Phosphoenolpyruvate
Carbohydrate Metabolism
Cyclic AMP
Adenosine

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Rabbit brain phosphofructokinase. Comparison of regulatory properties with those of other phosphofructokinase isozymes. / Tsai, M. Y.; Kemp, R. G.

In: Journal of Biological Chemistry, Vol. 249, No. 20, 01.12.1974, p. 6590-6596.

Research output: Contribution to journalArticle

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abstract = "A procedure was developed for the purification of phosphofructokinase from rabbit brain extracts. The enzyme was purified more than 100 fold with a yield of greater than 50{\%}. It was found to have the same distribution of multiple molecular forms as found in the total brain extract, including one molecular species that is different from the more extensively studied muscle (heart) and liver (erythrocyte) isozymes. The regulatory properties of brain phosphofructokinase were compared with those of muscle and liver phosphofructokinases at pH 7.1 and 7.4. All 3 enzymes were inhibited less at pH 7.4 than at 7.1. At pH 7.1 the sensitivity to ATP as an inhibitor decreased in the order liver > muscle > brain. At pH 7.4, the relative sensitivities of brain and muscle phosphofructokinases were reversed with the brain enzyme being inhibited at slightly lower concentrations of ATP. Other inhibitors acted synergistically with ATP. Sensitivity to inhibition by 3 phosphoglycerate and phosphoenolpyruvate decreased in the order muscle > brain > liver. With 2,3 diphosphoglycerate as an inhibitor, the sensitivity decreased in the order liver > muscle > brain. Muscle phosphofructokinase was the most sensitive to citrate inhibition. Creatine phosphate, a potent inhibitor of muscle phosphofructokinase, was completely ineffective as an inhibitor of liver and brain phosphofructokinase. The actions of activators were evaluated at pH 7.4 in the presence of inhibitory concentrations of ATP. The 3 enzymes were almost equally responsive to the deinhibiting action of AMP and cyclic adenosine 3':5' monophosphate, although the enzyme from liver requires slightly higher concentrations. Muscle phosphofructokinase was the least sensitive to deinhibition by inorganic phosphate. The results are discussed with respect to the varying modes of carbohydrate metabolism in the different tissues taking into consideration the tissue concentrations of the effectors and the variations of concentration in different physiological states. The enzymes from brain and liver are apparently chiefly controlled by the relative amounts of the adenine nucleotides and inorganic phosphate. An exception may be the erythrocyte, which has the same isozyme that is present in liver. Here, 2,3 diphosphoglycerate may play an important role in regulation. Muscle phosphofructokinase also responds to varying levels of adenine nucleotides but in vivo it is most likely that the principal regulatory effectors are creatine phosphate, citrate, and possibly, 3 phosphoglycerate.",
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