Open AccessIsolation and characterization of tetrahydropterin oxidation products generated in the tyrosine 3‐monooxygenase (tyrosine hydroxylase) reaction
Author(s) -
HAAVIK Jan,
FLATMARK Torgeir
Publication year - 1987
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1987.tb13381.x
Subject(s) - chemistry , monooxygenase , tyrosine 3 monooxygenase , tyrosine , phenylalanine hydroxylase , stereochemistry , tetrahydrobiopterin , substrate (aquarium) , cofactor , phenylalanine , enzyme , biochemistry , tyrosine hydroxylase , amino acid , cytochrome p450 , biology , ecology
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 (6 R )‐tetrahydrobiopterin, (6 RS )‐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.