1. Cyclic AMP-stimulated protein kinase activity phosphorylating intrinsic substrates in preparations of synaptic-membrane fragments from ox cerebral cortex was examined in relation to (a) the content of membrane-bound Ca2+in the preparations and (b) added Ca2+in the assay medium. 2. Centrifugal washing of synaptic-membrane fragments with buffered ethane dioxybis(ethylamine)tetra-acetate solutions decreased bound Ca2+from 2.8±0.4 (s.d.) to 0.9±0.3nmol/mg of protein. In washed preparations basal protein kinase activity was increased by about 40% and the cyclic AMP-stimulated activity by about 15%. Addition of Ca2+in the concentration range 5–50μm to the assay medium progressively inhibited the kinase activity of the washed preparations; in this range of Ca2+concentration the basal activity was inhibited more than the stimulated activity. 3. In unwashed preparations concentrations of Ca2+above 100μm inhibited the cyclic AMP-stimulated activity more than the basal activity. 4. The inhibitory effect of several concentrations of Ca2+was examined in relation to cyclic AMP concentration; no evidence for competition between Ca2+and cyclic AMP for a site on the enzyme was observed.
Protein kinase activity stimulated by adenosine 3′:5′-cyclic monophosphate in synaptic-membrane fragments from ox brain. Inhibition of intrinsic activity by free and membrane-bound calcium ions
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Malcolm Weller, Richard Rodnight; Protein kinase activity stimulated by adenosine 3′:5′-cyclic monophosphate in synaptic-membrane fragments from ox brain. Inhibition of intrinsic activity by free and membrane-bound calcium ions. Biochem J 1 September 1974; 142 (3): 605–609. doi: https://doi.org/10.1042/bj1420605
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