Macrophage oxidation of L-arginine to nitric oxide, nitrite, and nitrate. Tetrahydrobiopterin is required as a cofactor.
Macrophage oxidation of L-arginine to nitric oxide, nitrite, and nitrate. Tetrahydrobiopterin is required as a cofactor.
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发表时间：
1989-11
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通讯作者：
M. Tayeh;M. Marletta
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作者：
M. Tayeh;M. Marletta
最近在活化的巨噬细胞中表达的一种新的代谢途径的特征涉及作为中间体的一氧化氮（'N = O）的形成。形成的'N = O分解为亚硝酸盐（NO2-）和硝酸盐（NO3-）。该反应的底物是氨基酸精氨酸，其在胍基氮处被氧化以产生瓜氨酸作为反应的另一产物。本文报道的研究表明，在Sephadex G-25柱上层析后，100，000 x g上清液中所含的这种不寻常的氧化反应的活性丧失。一个小分子辅因子被认为是恢复活性所必需的。添加（6 R）-四氢生物蝶呤（BH 4）和NADPH导致该脱盐蛋白质的活性完全恢复。使用高效液相色谱和电化学检测对巨噬细胞提取物进行的分析表明，在巨噬细胞上清液中，BH 4的浓度为17 pmol/10（6）个细胞或2.1 μ M。仅存在（6 R）-异构体。随着BH 4和NADPH的加入，精氨酸的损失等于形成的NO2-、NO3-和瓜氨酸。在NADPH相对于BH 4的亚化学计量水平下，精氨酸的损失大于反应终产物的形成。与结果一致的反应途径的方案涉及精氨酸的N-羟基化作为初始步骤。BH 4参与这种类型的氧化化学反应与以前对这种辅因子的表征一致。
A new metabolic pathway characterized recently that is expressed in activated macrophages involves the formation of nitric oxide ('N = O) as an intermediate. The 'N = O formed decomposes to nitrite (NO2-) and nitrate (NO3-). The substrate for the reaction is the amino acid arginine which is oxidized at the guanido nitrogen to yield citrulline as the other product of the reaction. The studies reported here show that the activity for this unusual oxidation reaction which is contained in the 100,000 x g supernatant was lost after desalting on a Sephadex G-25 column. A small molecule cofactor was found to be required for the restoration of activity. The addition of (6R)-tetrahydrobiopterin (BH4) and NADPH led to complete recovery of activity in this desalted protein. Analysis of macrophage cell extracts, using high performance liquid chromatography with electrochemical detection, showed that BH4 was present at 17 pmol/10(6) cells or 2.1 microM in macrophage supernatant. Only the (6R)-isomer was present. With the addition of BH4 and NADPH, there was loss of arginine that was equal to the NO2-, NO3-, and citrulline formed. With substoichiometric levels of NADPH relative to BH4, the loss of arginine was greater than the formation of the end products of the reaction. A scheme for the reaction pathway consistent with the results involves N-hydroxylation of arginine as the initial step. The participation of BH4 in this type of oxidative chemistry is consistent with previous characterizations of this co-factor.