TYROSINE HYDROXYLATION CATALYZED BY MAMMALIAN TYROSINASE - IMPROVED METHOD OF ASSAY
TYROSINE HYDROXYLATION CATALYZED BY MAMMALIAN TYROSINASE - IMPROVED METHOD OF ASSAY
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DOI：
10.1016/0006-291x(64)90359-6
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发表时间：
1964-01-01
影响因子：
3.1
通讯作者：
POMERANTZ, SH
中科院分区：
文献类型：
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作者：
POMERANTZ, SH
本文报道了用L-酪氨酸-3，5-T研究酪氨酸酶催化的起始步骤：L-酪氨酸-3，5T+AH2+O2[前进箭头]3，4-二羟基苯丙氨酸-5T+TOH+A。水中氚的释放率与羟化速率成正比。可以很容易地比较不同的还原剂，并灵敏地确定初始速度。并将该方法与过量抗坏血酸蓄积多巴的方法进行了比较。这些实验表明：(1)哺乳动物酪氨酸酶的酪氨酸羟化需要氢供体的存在；(2)多巴是最有效的还原剂，但还原的蝶啶对苯丙氨酸羟化有效(Kaufman，1959)，对肾上腺和脑酪氨酸羟化酶有效(Nagatsu等人，1964)；(3)抗坏血酸，即使在高浓度，也不能消除酪氨酸与氧反应的滞后；(4)多巴可以作为酪氨酸羟化的竞争性抑制剂；(5)在催化量的多巴存在下，酪氨酸在高于约8×10~(-4)M的浓度时表现出明显的底物抑制；(6)很少或没有氚的速率效应。
This paper reports the use of L-tyrosine-3,5-T for studying the initial step catalyzed by tyrosinase: L-tyrosine-3,5T+AH2+O2[forward arrow]3,4-dihydroxy-phenylalanine -5T+TOH+A. The rate of tritium released as water is directly proportional to the rate of hydroxylation. Different reducing agents can be readily compared and initial rates sensitively determined. The method was compared with the technique of using excess ascorbate to accumulate dopa. These experiments indicate that (1) tyrosine hydroxylation by mammalian tyrosinase requires the presence of a hydrogen donor; (2) dopa is the most efficient reducing agent for this purpose, but a reduced pteridine, effective in phenylalanine hydroxylation (Kaufman, 1959) and with adrenal and brain tyrosine hydroxylase (Nagatsu et al., 1964), is effective at higher concentration; (3) ascorbate, even at high concentration, does not eliminate the lag in reaction of tyrosine with oxygen; (4) dopa can act as a competitive inhibitor of tyrosine hydroxylation; (5) tyrosine exhibits an apparent substrate inhibition at concentrations higher than about 8 x 10-4 M in the presence of catalytic quantities of dopa; (6) there is little or no tritium rate effect.