Autophosphorylation of Ca2+/calmodulin-dependent protein kinase II. Effects on total and Ca2+-independent activities and kinetic parameters.

Y. Hashimoto, C. M. Schworer, R. J. Colbran, Thomas Soderling

Research output: Contribution to journalArticle

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Abstract

Incubation of purified rat brain Ca2+/calmodulin-dependent protein kinase II for 2 min in the presence of Ca2+, calmodulin (CaM), Mg2+, and ATP converted the kinase from a completely Ca2+-dependent kinase to a substantially Ca2+-independent form with little loss of total activity. Subsequent addition of EGTA to the autophosphorylation reaction enhanced further autophosphorylation of the kinase which was associated with a suppression of total kinase activity to the Ca2+-independent value. Protein phosphatase 1 rapidly increased the suppressed total activity back to the control value and slowly decreased the Ca2+-independent activity. Kinetic analysis showed that the kinase not previously autophosphorylated had a Km for the synthetic peptide syntide-2 of 7 microM and Vmax of 9.8 mumol/min/mg when assayed in the presence of Ca2+ and CaM. The partially Ca2+-independent species, assayed in the presence of EGTA, had a Km of 21 microM and Vmax of 6.0. In the presence of Ca2+ and CaM the Km decreased and the Vmax increased to approximately control nonphosphorylated values. The completely Ca2+-independent form generated by sequential autophosphorylation first in the presence of Ca2+ and then EGTA had similar kinetic parameters to the partially independent species when assayed in the presence of EGTA, but addition of Ca2+ and CaM (up to 1 mg/ml) had little effect. These results suggest that separate autophosphorylation sites in the Ca2+/CaM-dependent protein kinase II are associated with formation of Ca2+-independent activity and suppression of total activity.

Original languageEnglish (US)
Pages (from-to)8051-8055
Number of pages5
JournalJournal of Biological Chemistry
Volume262
Issue number17
StatePublished - Jun 15 1987
Externally publishedYes

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Calcium-Calmodulin-Dependent Protein Kinase Type 2
Kinetic parameters
Egtazic Acid
Calmodulin
Phosphotransferases
syntide-2
Protein Phosphatase 1
Rats
Brain
Adenosine Triphosphate
Peptides
Kinetics

ASJC Scopus subject areas

  • Biochemistry

Cite this

Autophosphorylation of Ca2+/calmodulin-dependent protein kinase II. Effects on total and Ca2+-independent activities and kinetic parameters. / Hashimoto, Y.; Schworer, C. M.; Colbran, R. J.; Soderling, Thomas.

In: Journal of Biological Chemistry, Vol. 262, No. 17, 15.06.1987, p. 8051-8055.

Research output: Contribution to journalArticle

Hashimoto, Y. ; Schworer, C. M. ; Colbran, R. J. ; Soderling, Thomas. / Autophosphorylation of Ca2+/calmodulin-dependent protein kinase II. Effects on total and Ca2+-independent activities and kinetic parameters. In: Journal of Biological Chemistry. 1987 ; Vol. 262, No. 17. pp. 8051-8055.
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