TY - JOUR T1 - Activation of Tryptophan Hydroxylase by Adenosine Triphosphate, Magnesium, and Calcium JF - Molecular Pharmacology JO - Mol Pharmacol SP - 99 LP - 110 VL - 14 IS - 1 AU - MICHEL HAMON AU - SYLVIE BOURGOIN AU - F. HÉRY AU - G. SIMONNET Y1 - 1978/01/01 UR - http://molpharm.aspetjournals.org/content/14/1/99.abstract N2 - The addition of 0.5 mM ATP and 5 mM Mg2+ to a crude extract (35,000 x g supernatant) of a semipurified preparation (obtained by ammonium sulfate fractionation and Sephadex G-200 gel filtration) of tryptophan hydroxylase from rat brain stem induced a 40-60% increase in this enzymatic activity. The tryptophan hydroxylase remained in an activated state upon the removal of ATP and Mg2+ by filtration through a Sephadex G-25 column. The activation probably involved a phosphorylation reaction, since adenosine, a well-known protein kinase inhibitor, reduced the effects of ATP and Mg2+. In addition, when ATP was replaced by 5'-adenylylimidodiphosphate in the activating mixture, the activity of tryptophan hydroxylase remained unchanged, indicating that the terminal phosphate of ATP was required in the activating process. Although exogenous cyclic 3',5'-AMP-dependent protein kinase was able to increase further the activating effect of ATP and Mg2+ under some conditions (notably on the semipurified tryptophan hydroxylase), cyclic AMP failed to exert any effect on the enzyme in a crude extract. By contrast, when Ca2+ was included in the assay mixture, the stimulatory effect of ATP and Mg2+ on tryptophan hydroxylase activity was enhanced whatever the enzyme source. This effect of Ca2+ was maximal (25-30%) with a 10 µM concentration of the cation. These results strongly suggest that a Ca2+-dependent protein kinase may be involved in the activation of tryptophan hydroxylase induced by phosphorylating conditions. Kinetic analyses of tryptophan hydroxylase revealed that the apparent affinities of this enzyme for both tryptophan and DL-6-methyl-5,6,7,8-tetrahydropterin were increased under phosphorylating conditions. ACKNOWLEDGMENTS The authors with to thank Dr. J. Glowinski for continuous advice and encouragement. ER -