DOI: 10.1111/j.1471-4159.1980.tb11262.x. Purification and Some Characteristics of an ACTH‐Sensitive Protein Kinase and Its Substrate Protein in Rat Brain Membranes

Purification and Some Characteristics of an ACTH‐Sensitive Protein Kinase and Its Substrate Protein in Rat Brain Membranes

10.1111/j.1471-4159.1980.tb11262.x

Crossref journal-article

Wiley

Journal of Neurochemistry (311)

Abstract

Abstract: ACTH inhibits the phosphorylation of a rat brain membrane‐bound protein (B‐50). Both the protein kinase and the substrate protein could be extracted from the membranes by means of treatment with Triton X‐100 in 75 mM‐KCl. Using column chromatography over DEAE‐cellulose and ammonium sulphate precipitation a protein fraction (ASP 55–80) enriched in endogenous B‐50 phosphorylating activity was obtained. The time course of the endogenous phosphorylation of B‐50 in this fraction showed a linear incorporation with time for at least 10 min and reached an estimated maximal incorporation of 0.65 mol P/mol B‐50 after 60 min. The inhibition by ACTH1_24 of the B‐50 protein kinase in ASP 55–80 was dose‐dependent; the half‐maximal effective concentration was 5 × 10−6 M, being 10 to 50 times lower as compared with intact synaptic plasma membranes (SPM). cAMP, cGMP and various endor‐phins had no effect on the B‐50 protein kinase. The B‐50 protein kinase required both magnesium and calcium for optimal activity. Using two‐dimensional electrophoresis on polyacrylamide slab gels the B‐50 protein kinase and the B‐50 protein could be identified and purified. The isoelectric point (IEP) of the kinase is 5.5 and the apparent molecular weight 70,000, whereas the IEP of the substrate protein B‐50 is 4.5 and the apparent molecular weight 48,000. Amino acid analysis on microgram quantities of purified kinase and B‐50 protein revealed basic/acidic amino acid ratios in agreement with the respective lEP's. It is speculated that the inhibition of B‐50 protein kinase may be related to known modulatory effects of ACTH and related peptides on certain types of neurotransmission and behaviour.

Bibliography

Zwiers, H., Schotman, P., & Gispen, W. H. (1980). Purification and Some Characteristics of an ACTHâSensitive Protein Kinase and Its Substrate Protein in Rat Brain Membranes. Journal of Neurochemistry, 34(6), 1689â1699. Portico.

Authors 3

H. Zwiers (first)
P. Schotman (additional)
W. H. Gispen (additional)

References 25 Referenced 175

{'key': 'e_1_2_2_2_1', 'first-page': '1281', 'article-title': 'Neuropeptides and protein phosphorylation in brain membranes', 'volume': '4', 'author': 'Davis L. G.', 'year': '1978', 'journal-title': 'Soc. Neurosc.'} / Soc. Neurosc. / Neuropeptides and protein phosphorylation in brain membranes by Davis L. G. (1978)
10.1016/0006-291X(76)90828-7
10.1073/pnas.76.4.1838
10.1016/S0021-9258(17)34272-2 / J. Biol. Chem. / Inhibition of cyclic AMP‐dependent protein kinase by analogues of a synthetic peptide substrate by Feramisco J. R. (1978)
10.1016/0024-3205(75)90103-4
10.1007/978-1-4684-3503-0_11
10.1016/0304-4165(78)90023-5
10.1073/pnas.73.4.1038
10.1016/S0021-9258(19)52451-6
10.1111/j.1471-4159.1978.tb12459.x
{'key': 'e_1_2_2_12_1', 'first-page': '96', 'article-title': 'The effect of morphine on the endogenous phosphorylation of synaptic plasma membrane proteins of rat striatum', 'volume': '208', 'author': "O'Callaghan J. P.", 'year': '1979', 'journal-title': 'J. Pharmacol. Exp. Ther.'} / J. Pharmacol. Exp. Ther. / The effect of morphine on the endogenous phosphorylation of synaptic plasma membrane proteins of rat striatum by O'Callaghan J. P. (1979)
{'key': 'e_1_2_2_13_1', 'volume-title': 'Ahstr. 7th Int. Meet', 'author': 'Oestreicher A. B.', 'year': '1979'} / Ahstr. 7th Int. Meet by Oestreicher A. B. (1979)
10.1016/S0021-9673(00)97051-1
10.1073/pnas.76.5.2475
10.1111/j.1600-0773.1973.tb01477.x
10.1016/S0021-9258(17)40170-0 / j. Biol. Chem. / Adenosine 3′, 5′‐monophos‐phate‐regulated phosphoprotein system of neuronal membranes. I. Solubilization, purification and some properties of an endogenous phosphoprotein by Ueda T. (1977)
10.1016/S0021-9258(17)33678-5 / J. Biol. Chem. / Differences in properties of cytosol and membrane‐derived protein kinases by Uno I. (1976)
10.1016/S0021-9258(17)40171-2 / J. Biol. Chem. / Adenosine 3′, 5′‐monophosphate‐regulated phosphoprotein system of neuronal membranes. II. Solubilization, purification and some properties of an endogenous adenosine 3′, 5′‐monophosphate‐dependent protein kinase by Uno I. (1977)
10.1007/978-1-4899-7321-4_14
10.1007/BF00966326
WinterA. EkK.andAnderssonU. B.(1977)Analytical elec‐trofocussing in thin‐layer polyacrylamide gels.L.K.B. Application Note250.
10.1007/BF00965607
{'key': 'e_1_2_2_24_1', 'first-page': '267', 'volume-title': 'Mechanisms, Regulation and Special Function of Protein Synthesis in the Brain', 'author': 'Zwiers H.', 'year': '1977'} / Mechanisms, Regulation and Special Function of Protein Synthesis in the Brain by Zwiers H. (1977)
10.1007/BF00966327
10.1111/j.1471-4159.1979.tb11726.x

Dates

Type	When
Created	18 years, 10 months ago (Oct. 5, 2006, 6:25 a.m.)
Deposited	1 year, 10 months ago (Nov. 4, 2023, 5:05 a.m.)
Indexed	59 minutes ago (Sept. 4, 2025, 10:17 a.m.)
Issued	45 years, 3 months ago (June 1, 1980)
Published	45 years, 3 months ago (June 1, 1980)
Published Online	18 years, 10 months ago (Oct. 5, 2006)
Published Print	45 years, 3 months ago (June 1, 1980)

Funders 0

None

BibTeX

@article{Zwiers_1980, title={Purification and Some Characteristics of an ACTH‐Sensitive Protein Kinase and Its Substrate Protein in Rat Brain Membranes}, volume={34}, ISSN={1471-4159}, url={http://dx.doi.org/10.1111/j.1471-4159.1980.tb11262.x}, DOI={10.1111/j.1471-4159.1980.tb11262.x}, number={6}, journal={Journal of Neurochemistry}, publisher={Wiley}, author={Zwiers, H. and Schotman, P. and Gispen, W. H.}, year={1980}, month=jun, pages={1689–1699} }

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