Regulation of eNOS-derived superoxide by endogenous methylarginines
Regulation of eNOS-derived superoxide by endogenous methylarginines
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DOI：
10.1021/bi702377a
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发表时间：
2008-07-08
期刊：
影响因子：
2.9
通讯作者：
Cardounel, Arturo J.
中科院分区：
文献类型：
--
作者：
Druhan, Lawrence J.;Forbes, Scott P.;Cardounel, Arturo J.
内源性甲基精氨酸、不对称二甲基精氨酸（ADMA）和n - g -单甲基- l-精氨酸（L-NMMA）调节内皮NO合成酶（eNOS）产生一氧化氮（NO）。在四氢生物蝶素（BH4）耗竭条件下，eNOS也生成o中心点(2)-；然而，甲基精氨酸对enos衍生的o -中心点(2)-生成的影响尚不清楚。因此，利用电子顺磁共振自旋捕获技术，我们测量了在BH4耗损条件下ADMA和L-NMMA对eNOS生成o -中心点(2)-的剂量依赖性效应。在没有BH4的情况下，ADMA剂量依赖性地增加了nos衍生中心点O-2（-）的产生，在100 μ M ADMA时最大增幅为151%。L-NMMA也呈剂量依赖性地增加nos衍生的中心点O-2(-)，但幅度较小，在100 μ M L-NMMA时增加102%。此外，天然底物l -精氨酸也增加了enos衍生的中心点O-2(-)，表现出与ADMA相似的增强程度。eNOS对NADPH消耗的测量表明，l -精氨酸或甲基精氨酸的结合都增加了NADPH氧化的速度。分光光度法研究表明，就像l -精氨酸和L-NMMA一样，ADMA的结合将eNOS血红素转移到高自旋状态，表明血红素氧化还原电位更正，从而增强了从还原酶到加氧酶位点的电子转移。这些结果表明，甲基精氨酸可以深刻地改变eNOS中NO和中心点O-2（-）生成的平衡。这些观察结果对于l -精氨酸和甲基精氨酸- nos抑制剂在疾病治疗中的应用具有重要意义。
The endogenous methylarginines, asymmetric dimethylarginine (ADMA) and N-G-monomethyl-L-arginine (L-NMMA) regulate nitric oxide (NO) production from endothelial NO synthase (eNOS). Under conditions of tetrahydrobiopterin (BH4) depletion eNOS also generates O-center dot(2)-; however, the effects of methylarginines on eNOS-derived O-center dot(2)- generation are poorly understood. Therefore, using electron paramagnetic resonance spin trapping techniques we measured the dose-dependent effects of ADMA and L-NMMA on O-center dot(2)- production from eNOS under conditions of BH4 depletion. In the absence of BH4, ADMA dose-dependently increased NOS-derived center dot O-2(-) generation, with a maximal increase of 151% at 100 mu M ADMA. L-NMMA also dose-dependently increased NOS-derived center dot O-2(-), but to a lesser extent, demonstrating a 102% increase at 100 mu M L-NMMA. Moreover, the native substrate L-arginine also increased eNOS-derived center dot O-2(-), exhibiting a similar degree of enhancement as that observed with ADMA. Measurements of NADPH consumption from eNOS demonstrated that binding of either L-arginine or methylarginines increased the rate of NADPH oxidation. Spectrophotometric studies suggest, just as for L-arginine and L-NMMA, the binding of ADMA shifts the eNOS heme to the high-spin state, indicative Of a more positive heme redox potential, enabling enhanced electron transfer from the reductase to the oxygenase site. These results demonstrate that the methylarginines can profoundly shift the balance of NO and center dot O-2(-) generation from eNOS. These observations have important implications with regard to the therapeutic use of L-arginine and the methylarginine-NOS inhibitors in the treatment of disease.