Isolation and characterization of tetrahydropterin oxidation products generated in the tyrosine 3-monooxygenase (tyrosine hydroxylase) reaction.
The catalytic mechanism of tyrosine 3-monooxygenase (tyrosine hydroxylase, EC 1.14.16.2), isolated from the cytosolic fraction of bovine adrenal medulla, was studied by new techniques of product isolation and characterization. Using either (6R)-tetrahydrobiopterin, (6RS)-tetrahydroneopterin, or 6-methyl-tetrahydropterin, as the cofactors, three enzymatic oxidation products could be isolated and identified from the reaction mixture by high-performance liquid chromatography and rapid-scanning spectroscopy: (a) the 4a-hydroxy derivatives, (b) the quinonoid dihydropterins, and (c) the stable 7,8-dihydropterins. Stopped-flow spectroscopy revealed that the formation of the 4a-hydroxy-tetrahydropterins preceded the formation of the quinonoid forms with both L-tyrosine and L-phenylalanine as the substrate. The formations of 4a-hydroxy-tetrahydropterins and hydroxylated amino acids were tightly coupled as recently shown in the phenylalanine 4-monooxygenase reaction [Haavik, J., Døskeland, A. P. & Flatmark, T. (1986) Eur. J. Biochem. 160, 1-8]. No detectable carbinolamine dehydratase activity was present in the enzyme preparation.
D000313:Adrenal Medulla / Q000201:enzymology / Q000378:metabolism; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000201:enzymology; D010084:Oxidation-Reduction; D011622:Pterins / Q000378:metabolism*; D014446:Tyrosine 3-Monooxygenase / Q000302:isolation & purification*
D011622:Pterins; C011552:tetrahydropterin; D014446:Tyrosine 3-Monooxygenase