Premium
Ca 2+ Sensitivity of Ca 2+ ‐Dependent Protein Kinase Activities Toward Intrinsic Proteins in Synaptosomal Membrane Fragments from Rat Cerebral Tissue
Author(s) -
Gower Hilary,
Rodnight Richard,
Brammer Michael J.
Publication year - 1986
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1986.tb12988.x
Subject(s) - chemistry , biophysics , membrane , protein kinase a , biochemistry , kinase , biology
The Ca 2+ and calmodulin sensitivity of endogenous protein kinase activity in synaptosomal membrane fragments from rat brain was studied in medium containing Ca 2+ plus EGTA using a modified computer programme to calculate free Ca 2+ concentrations that took into account the effect of all competing cations and chelators. The Ca 2+ ‐dependent phosphorylation of 10 major polypeptide acceptors with M r values ranging from 50 to 360 kilodaltons required calmodulin in reactions that were all equally sensitive to Ca 2+ ; half‐maximal phosphorylation required a free Ca 2+ concentration of 45 n M and maximal phosphorylation ∼110 n M . The significance of these values in relation to published data on the intracellular concentration of free Ca 2+ in the nervous system is discussed. One acceptor of 45 kilodaltons was phosphorylated in a Ca 2+ ‐dependent reaction that did not require calmodulin. This polypeptide appeared to correspond to the B‐50 protein, an established substrate of the lipid‐dependent protein kinase C. Further study of this phosphorylating system showed that the reaction was only independent of calmodulin at saturating concentrations of Ca 2+ ; at subsaturating concentrations (in the range 50–130 n M ), a small but significant stimulation of the enzyme by calmodulin was demonstrated. The possible significance of this finding is discussed.