FORMATION OF FREE NITRIC-OXIDE FROM L-ARGININE BY NITRIC-OXIDE SYNTHASE - DIRECT ENHANCEMENT OF GENERATION BY SUPEROXIDE-DISMUTASE
FORMATION OF FREE NITRIC-OXIDE FROM L-ARGININE BY NITRIC-OXIDE SYNTHASE - DIRECT ENHANCEMENT OF GENERATION BY SUPEROXIDE-DISMUTASE
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DOI：
10.1073/pnas.91.23.10992
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发表时间：
1994-11-08
影响因子：
11.1
通讯作者：
IGNARRO, LJ
中科院分区：
文献类型：
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作者：
HOBBS, AJ;FUKUTO, JM;IGNARRO, LJ
尽管一氧化氮 (NO) 似乎是 NO 合酶 (NOS; EC 1.14.13.39) 催化的 L-精氨酸氧化产物之一，但过去在无细胞酶测定中测量 NO 的研究并不是基于游离 NO 分子的直接检测。相反，测定依赖于稳定的NO氧化产物亚硝酸盐和硝酸盐的间接测量以及NO的间接作用，例如鸟苷酸环化酶激活和氧合血红蛋白氧化。利用特定的化学发光测定，我们在此报告，由诱导型巨噬细胞和组成型神经元 NOS 催化的 L-精氨酸氧化的气态产物，其物理化学性质与真正的 NO 没有区别。 NOS 形成的 NO 气体依赖于 L-精氨酸、NADPH 和氧气，并受到 N-G-甲基-L-精氨酸和氰化物阴离子的抑制。超氧化物歧化酶 (SOD) 通过与超氧阴离子歧化或 NOS 活化无关的机制引起游离 NO 产生的显着、浓度依赖性增加。这些观察结果表明，游离NO是NOS催化的L-精氨酸氧化的结果，并且SOD增强NO的生成而不直接影响NO本身。 SOD 似乎能引发一种新的生物作用，可能会加速 L-精氨酸-NO 途径中的中间体（例如硝酰基 (HNO)）向 NO 的转化。
Although nitric oxide (NO) appears to be on of the oxidation products of L-arginine catalyzed by NO synthase (NOS; EC 1.14.13.39), past studies on the measurement of NO in cell-free, enzymatic assays have not been based on the direct detection of the free NO molecule. Instead, assays have relied on indirect measurements of the stable NO oxidation products nitrite and nitrate and on indirect actions of NO such as guanylate cyclase activation and oxyhemoglobin oxidation. Utilizing a specific chemiluminescence assay, we report here that the gaseous product of L-arginine oxidation, catalyzed by both inducible macrophage and constitutive neuronal NOS, is indistinguishable from authentic NO on the basis of their physicochemical properties. NO gas formation by NOS was dependent on L-arginine, NADPH, and oxygen and inhibited by N-G-methyl-L-arginine and cyanide anion. Superoxide dismutase (SOD) caused a marked, concentration-dependent increase in the production of free NO by mechanisms that were unrelated to the dismutation of superoxide anion or activation of NOS. These observations indicate that free NO is formed as a result of NOS-catalyzed L-arginine oxidation and that SOD enhances the generation of NO without directly affecting NO itself. SOD appears to elicit a novel biological action, perhaps accelerating the conversion of an intermediate in the L-arginine-NO pathway such as nitroxyl (HNO) to NO.