JBC, Vol. 252, Issue 9, 3007-3011, May, 1977
Ionic inhibition of catalytic phosphorylation of histone by bovine brain protein kinase
G. W. Moll Jr and E. T. Kaiser
The effects of various ions commonly found in protein kinase assays upon
the rate of histone phosphorylation catalyzed by the highly purified bovine
brain enzyme, protein kinase I, have been investigated. Sodium, potassium,
and magnesium were found to inhibit histone phosphorylation by protein
kinase I in a similar manner. The degree of inhibition by any of these
cations was demonstrated to be directly proportional to the square root of
the ionic strength of the assay medium. The relationship between the ionic
strength of the assay medium and the rate of histone phosphorylation
catalyzed by protein kinase I was employed to correct the rate of histone
phosphorylation at various magnesium acetate concentrations to a standard
ionic strength. When this was done an analysis of the previously postulated
rate law for histone phosphorylation c atalyzed by protein kinase I gave a
binding constant for the magnesium-ATP complex which was in agreement with
that expected for this complex on the basis of various binding constants
available in the literature. These results demonstrate that it is
unnecessary to postulate a specific ion inhibition process for protein
kinase I by the ions employed in this study. They also support the
reasonable assumption that magnesium ion binds to ATP at or prior to the
rate-determining step in histone phosphorylation catalyzed by protein
kinase I. The expression developed in this paper for the effect of ionic
strength upon protein kinase I activity can now be used to correct activity
measurements made under various assay conditions to a standard assay state,
allowing facile comparisons of kinetic data. It should be possible to
develop similar expressions for other protein kinases and substrates to
permit useful interpretation of kinetic data.
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