Further studies on tryptophan hydroxylase from neoplastic murine mast cells.
Tryptophan hydroxylase (tryptophan, tetrahydropteridine: oxygen oxidoreductase (5-hydroxylating) EC 1.14.16.4) purified from the neoplastic murine mast cells by hydroxylapatite chromatography following ammonium sulfate fractionation showed maximum activity at pH 6.0 in the presence of 2-mercaptoethanol, 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetra-hydropteridine and Fe2+, and pH 7.6 to 8.0 in the absence of addED Fe2+. The Km values were 38.5 muM and 22.2 muM for tryptophan, 298 muM and 204 muM for 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetra-hydropteridine, and 6.45% for oxygen in either presence or absence of added Fe-2+, respectively. From kinetic data the reaction mechanism of tryptophan hydroxylation appears to be of the sequential, rather than the ping-pong, type. Tryptophan hydroxylase from mast cells was considerably inhibited by o-phenanthroline like phenylalanine hydroxylase as well as tyrosine hydroxylase from other sources, and its Ki was between 1.2 muM and 4.53 muM. It was found that the inhibition by o-phenanthroline was competitive with respect to both tryptophan and 2-amino-4-hydroxy-6,7-dimethyl-5,6,7,8-tetrahydropteridine, but not molecular oxygen under the assay conditions employed.
D000818:Animals; D001667:Binding, Competitive; D002374:Catalase; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007501:Iron / Q000378:metabolism; D007700:Kinetics; D012515:Mast-Cell Sarcoma / Q000201:enzymology*; D008623:Mercaptoethanol / Q000494:pharmacology; D051379:Mice; D008811:Mice, Inbred DBA; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009584:Nitrogen; D010100:Oxygen; D010618:Phenanthrolines / Q000494:pharmacology; D011621:Pteridines / Q000494:pharmacology; D014365:Tryptophan Hydroxylase / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*
D010618:Phenanthrolines; D011621:Pteridines; D008623:Mercaptoethanol; D007501:Iron; D006899:Mixed Function Oxygenases; D002374:Catalase; D014365:Tryptophan Hydroxylase; D009584:Nitrogen; D010100:Oxygen