Levine M, Morita K, Heldman E, Pollard H B
J Biol Chem. 1985 Dec 15;260(29):15598-603.
The effect of ascorbic acid on the conversion of dopamine to norepinephrine was investigated in isolated chromaffin granules from bovine adrenal medulla. Ascorbic acid was shown to double the rate of [3H]norepinephrine formation from [3H]dopamine, despite no demonstrable accumulation of ascorbic acid into chromaffin granules. The enhancement of norepinephrine biosynthesis by ascorbic acid was dependent on the external concentrations of dopamine and ascorbate. The apparent Km of the dopamine beta-hydroxylation system for external dopamine was approximately 20 microM in the presence or absence of ascorbic acid. However, the apparent maximum velocity of norepinephrine formation was nearly doubled in the presence of ascorbic acid. By contrast, the apparent Km and Vmax of dopamine uptake into chromaffin granules were not affected by ascorbic acid. Norepinephrine formation was increased by ascorbic acid when the concentration of ascorbate was 200 microM or higher; a concentration of 2 mM appeared to induce the maximal effect under the experimental conditions used here. The effect of ascorbic acid on conversion of dopamine to norepinephrine required Mg-ATP-dependent dopamine uptake into chromaffin granules. In contrast to ascorbic acid, other reducing agents such as NADH, glutathione, and homocysteine were unable to enhance norepinephrine biosynthesis. These data suggest that ascorbic acid provides reducing equivalents for hydroxylation of dopamine despite the lack of ascorbate accumulation into chromaffin granules. These findings imply the functional existence of an electron carrier system in the chromaffin granule which transfers electrons from external ascorbic acid for subsequent intragranular norepinephrine biosynthesis.
在从牛肾上腺髓质分离出的嗜铬颗粒中，研究了抗坏血酸对多巴胺转化为去甲肾上腺素的影响。结果显示，抗坏血酸能使[3H]多巴胺生成[3H]去甲肾上腺素的速率加倍，尽管未检测到抗坏血酸在嗜铬颗粒中的明显积累。抗坏血酸对去甲肾上腺素生物合成的增强作用取决于多巴胺和抗坏血酸盐的外部浓度。无论有无抗坏血酸，多巴胺β-羟化系统对外部多巴胺的表观Km约为20微摩尔。然而，在有抗坏血酸存在的情况下，去甲肾上腺素生成的表观最大速度几乎增加了一倍。相比之下，抗坏血酸对嗜铬颗粒摄取多巴胺的表观Km和Vmax没有影响。当抗坏血酸盐浓度为200微摩尔或更高时，抗坏血酸可增加去甲肾上腺素的生成；在此处使用的实验条件下，2毫摩尔的浓度似乎能诱导最大效应。抗坏血酸对多巴胺转化为去甲肾上腺素的作用需要Mg-ATP依赖的多巴胺摄取进入嗜铬颗粒。与抗坏血酸不同，其他还原剂如NADH、谷胱甘肽和同型半胱氨酸无法增强去甲肾上腺素的生物合成。这些数据表明，尽管抗坏血酸没有在嗜铬颗粒中积累，但它为多巴胺的羟化提供了还原当量。这些发现意味着嗜铬颗粒中存在一个电子载体系统，该系统从外部抗坏血酸转移电子，用于随后的颗粒内去甲肾上腺素生物合成。
