Properties of Protein Kinases in Brain Coated Vesicles

• Published in: Journal of biochemistry (Tokyo) Vol. 98; no. 1; pp. 63 - 68
• Main Authors: TAKAHASHI, Akira, USAMI, Mihoko, KADOTA, Tomoko, KADOTA, Ken
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01.01.1985
• Subjects: Adenosine Triphosphate - metabolism; Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; brain; Brain - enzymology; Cattle; Coated Pits, Cell-Membrane - metabolism; coated vesicles; Endosomes - metabolism; Enzymes and enzyme inhibitors; Fundamental and applied biological sciences. Psychology; Guanosine Triphosphate - metabolism; Heparin - pharmacology; Kinetics; Manganese - pharmacology; Membrane Proteins - metabolism; Phosvitin - metabolism; protein kinase; Protein Kinases - metabolism; Salts - pharmacology; Substrate Specificity; Transferases; Troponin - metabolism; Troponin T; Characterization; Coated vesicle; Bovine; Enzyme; Protein kinase; Transferase; Animal; Brain(Invertebrata); Catalyst activity; Physicochemical properties
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/oxfordjournals.jbchem.a135273

Coated vesicles prepared from bovine brain contained cyclic nucleotides- and Ca2+ calmodulin-independent protein kinases which in the presence of Mg2+ catalyzed the phosphorylation of an endogenous 48,000 Mr protein of coated vesicles (C-48), phosvitin and troponin T. Phosvitin was phosphorylated either in the presence of ATP or GTP. The phosphorylation of C-48, on the other hand, was specific for ATP. Heparin inhibited the phosphorylation of phosvitin but not that of C-48. Mn2+ inhibited the phosphorylation of phosvitin, while Mn2+ substituted for Mg2+ in the phosphorylation of C-48. When the coated vesicles were prepared in the presence of NaF, C-48 contained 2.5-2.8 mol of phosphate/mol. On incubation with Mg2+ and ATP, C-48 incorporated 1.2–1.6 mol of phosphate/mol. With C-48 as a substrate, the value of its apparent Km for ATP was 6 μM. With phosvitin as a substrate, the value of its apparent Km was 20 μM. The phosphorylated amino acid residues in the phosvitin were identified as serine and threonine. Phosphothreonine was detected in C-48. These results suggest that brain coated vesicles possess two different classes of protein kinase, a casein kinase II and C-48 kinase.