Glycogen synthase kinase-3β is a dual specificity kinase differentially regulated by tyrosine and serine/threonine phosphorylation

Q. May Wang, Carol J. Fiol, Anna De Paoli-Roach, Peter Roach

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Abstract

The enzyme glycogen synthase kinase-3 (GSK-3) has been implicated in the control of several metabolic enzymes and transcription factors in response to extracellular signals. In the past, the enzyme has been considered to be a protein Ser/Thr kinase although it was recently reported to contain Tyr(P) (Hughes, K., Nikolakaki, E., Plyte, S. E., Totty, N. F., and Woodgett, J. R. (1993) EMBO J. 12, 803-808). A cDNA encoding rabbit skeletal muscle GSK-3β was cloned and expressed in Escherichia coli as an active protein kinase, with apparent M(r) 46,000, capable of phosphorylating several known GSK-3 substrates. Recombinant GSK-3β autophosphorylated on Ser, Thr, and Tyr residues although the enzyme already contained Tyr(P) as judged by its recognition by anti-Tyr(P) antibodies. The net result of the autophosphorylation was a 3-5-fold reduction in enzyme activity. GSK-3α, purified from rabbit muscle, also underwent autophosphorylation but only on Ser and Thr residues. In this case, the autophosphorylation stabilized the enzyme activity compared with the control lacking ATP/Mg2+. Of several phosphatases tested, the λ-phage phosphatase was the most effective in dephosphorylating at Ser and Thr residues but did not dephosphorylate at Tyr residues. The action of the λ-phosphatase caused a reactivation of GSK-3β to ~80% of the starting activity. The protein tyrosine phosphatase PTP1B was able to dephosphorylate at Tyr residues leading to a reduction in enzyme activity. A truncated form of GSK-3β, apparent M(r) 40,000, had a significantly higher specific activity, was defective in autophosphorylation, and was not inactivated in the autophosphorylation reaction. We conclude that GSK-3β is a dual specificity protein kinase in the same sense as the mitogen-activated protein kinase/ERK family of enzymes. Phosphorylation at different residues differentially controls enzyme activity, Ser/Thr phosphorylation causing inactivation and Tyr phosphorylation resulting in increased activity.

Original languageEnglish
Pages (from-to)14566-14574
Number of pages9
JournalJournal of Biological Chemistry
Volume269
Issue number20
StatePublished - May 20 1994

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Glycogen Synthase Kinase 3
Phosphorylation
Threonine
Serine
Tyrosine
Phosphotransferases
Enzyme activity
Enzymes
Phosphoric Monoester Hydrolases
Protein Kinases
Muscle
Bacteriophages
Rabbits
Protein Tyrosine Phosphatases
Mitogen-Activated Protein Kinases
Escherichia coli
Transcription Factors
Complementary DNA
Adenosine Triphosphate
Skeletal Muscle

ASJC Scopus subject areas

  • Biochemistry

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Glycogen synthase kinase-3β is a dual specificity kinase differentially regulated by tyrosine and serine/threonine phosphorylation. / Wang, Q. May; Fiol, Carol J.; De Paoli-Roach, Anna; Roach, Peter.

In: Journal of Biological Chemistry, Vol. 269, No. 20, 20.05.1994, p. 14566-14574.

Research output: Contribution to journalArticle

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abstract = "The enzyme glycogen synthase kinase-3 (GSK-3) has been implicated in the control of several metabolic enzymes and transcription factors in response to extracellular signals. In the past, the enzyme has been considered to be a protein Ser/Thr kinase although it was recently reported to contain Tyr(P) (Hughes, K., Nikolakaki, E., Plyte, S. E., Totty, N. F., and Woodgett, J. R. (1993) EMBO J. 12, 803-808). A cDNA encoding rabbit skeletal muscle GSK-3β was cloned and expressed in Escherichia coli as an active protein kinase, with apparent M(r) 46,000, capable of phosphorylating several known GSK-3 substrates. Recombinant GSK-3β autophosphorylated on Ser, Thr, and Tyr residues although the enzyme already contained Tyr(P) as judged by its recognition by anti-Tyr(P) antibodies. The net result of the autophosphorylation was a 3-5-fold reduction in enzyme activity. GSK-3α, purified from rabbit muscle, also underwent autophosphorylation but only on Ser and Thr residues. In this case, the autophosphorylation stabilized the enzyme activity compared with the control lacking ATP/Mg2+. Of several phosphatases tested, the λ-phage phosphatase was the most effective in dephosphorylating at Ser and Thr residues but did not dephosphorylate at Tyr residues. The action of the λ-phosphatase caused a reactivation of GSK-3β to ~80{\%} of the starting activity. The protein tyrosine phosphatase PTP1B was able to dephosphorylate at Tyr residues leading to a reduction in enzyme activity. A truncated form of GSK-3β, apparent M(r) 40,000, had a significantly higher specific activity, was defective in autophosphorylation, and was not inactivated in the autophosphorylation reaction. We conclude that GSK-3β is a dual specificity protein kinase in the same sense as the mitogen-activated protein kinase/ERK family of enzymes. Phosphorylation at different residues differentially controls enzyme activity, Ser/Thr phosphorylation causing inactivation and Tyr phosphorylation resulting in increased activity.",
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