title: Tryptophan hydroxylase: purification by affinity chromatography on calmodulin-sepharose.
abstract: Tryptophan hydroxylase (EC 1.14.16.4; L-tryptophan, tetrahydropteridine: oxygen oxidoreductase (5-hydroxylating)) from rat mesencephalic tegmentum has been purified by sequential chromatography on Blue-Sepharose, DE-52, and calmodulin-Sepharose. The hydroxylase is excluded from Blue-Sepharose and is eluted from DE-52 with a step-wise NaCl gradient. Tryptophan hydroxylase binds to calmodulin-Sepharose in the presence of calcium and is eluted with either EGTA or calmodulin itself, but not with tryptophan. The purification scheme is rapid (5-6 h) and yields an enzyme with a specific activity of 225 nmol 5-HTP/mg min, representing a 400-fold purification with 7% recovery. The tryptophan hydroxylase preparation was judged to be > 95% pure using the present isolation procedure.
authors.affiliation: Department of Psychiatry and Behavioral Neurosciences, Wayne State University School of Medicine, MI 48201, USA.
authors.forename: C
authors.initials: C
authors.lastname: D'Sa
authors.forename: R
authors.initials: R
authors.lastname: Arthur
authors.forename: I
authors.initials: I
authors.lastname: Jennings
authors.forename: R G
authors.initials: RG
authors.lastname: Cotton
authors.forename: D M
authors.initials: DM
authors.lastname: Kuhn
mesh_terms: D000818:Animals; D001921:Brain / Q000201:enzymology; D002147:Calmodulin; D002846:Chromatography, Affinity / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D012685:Sepharose; D014365:Tryptophan Hydroxylase / Q000302:isolation & purification*
chemical_list: D002147:Calmodulin; D012685:Sepharose; D014365:Tryptophan Hydroxylase