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Properties of the 12‐ O ‐Tetradecanoylphorbol‐13‐Acetate‐Stimulated S6 Kinase from Rat Astroglial Cells
Author(s) -
ToruDelbauffe Danièle,
Gavaret JeanMichel,
Jacquemin Claude,
Matricon Carole,
Pomerance Martine,
Pierre Michel
Publication year - 1988
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.1471-4159.1988.tb01110.x
Subject(s) - 12 o tetradecanoylphorbol 13 acetate , enzyme , protein kinase a , biochemistry , chemistry , sedimentation coefficient , kinase , microbiology and biotechnology , phosphorylation , protein kinase c , biology , phorbol ester
The S6 kinase activity of astroglial cells in primary culture stimulated by 12‐ O ‐tetradecanoylphorbol‐13‐acetate (TPA) has been studied. This activity was eluted as a single peak at 0.15 M NaCl from a DEAE‐Sephacel column. The chromatography of this peak on phosphocellulose revealed an activity eluted at 0.15 M NaCl. This partially purified enzyme had a sedimentation coefficient of 3.7S; K m values were 2 × 10 −5 M for ATP and 10 −6 M for 40S ribosomal subunits. The optimal Mg 2+ concentration requirement was 2‐3 m M . Mn 2+ and Co 2+ could substitute for Mg 2+ (optimum concentrations 1.5 and 0.8 m M , respectively), but these cations were strong inhibitors in the presence of Mg 2+ . The enzyme was inhibited by N ‐ethylmaleimide, indicating that it contained thiol groups. This S6 kinase used ATP, but not GTP, as a phosphate donor, and exhibited great specificity for S6 as phosphate acceptor. Whole histones and protamine were slightly phosphorylated whereas phosvitin, histone H1. and surprisingly the peptide Arg‐Arg‐Leu‐Ser‐Ser‐Leu‐Arg‐Ala were not phosphorylated. The TPA‐stimulated S6 kinase resembles the insulin‐, fibroblast growth factor‐ and cyclic AMP‐stimulated enzymes, suggesting that several pathways might activate the same entity.