Regulation of inducible nitric oxide synthase by self-generated NO
Regulation of inducible nitric oxide synthase by self-generated NO
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DOI：
10.1021/bi010066m
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发表时间：
2001-06-12
期刊：
影响因子：
2.9
通讯作者：
Stuehr, DJ
中科院分区：
文献类型：
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作者：
Abu-Soud, HM;Ichimori, K;Stuehr, DJ
L-精氨酸合成 NO 时，小鼠诱导型一氧化氮合酶 (iNOS) 中会形成铁血红素 - 一氧化氮 (NO) 复合物。我们研究了其形成动力学、对催化活性的影响、对溶液 NO 浓度的依赖性以及对酶氧反应（表观 KmO(2)）的影响。血红素-NO 复合物的形成是双相的，并且在动力学上与 iNOS 中电子通量和催化的抑制有关。利用超氧化物生成系统清除 NO 的实验表明，血红素-NO 复合物形成的程度直接取决于反应溶液中达到的 NO 浓度。然而，即使溶液 NO 被完全清除，在 NO 合成过程中仍会形成一小部分血红素-NO 复合物 (20%)。内在血红素-NO 复合物的形成以及与溶液 NO 的积累相关的血红素-NO 复合物的形成，使 iNOS 的表观 KmO2 分别增加了 10 倍和 4 倍。总之，数据显示 iNOS 中血红素-NO 复合物的形成是由于内在的 NO 结合和溶液 NO 的平衡结合所致，当 NO 达到高纳摩尔至低微摩尔浓度时，后者占主导地位。这种行为将 iNOS 与其他 NOS 亚型区分开来，并表明其在生物环境中的活性和氧反应可能有更复杂的调节。
A ferric heme-nitric oxide (NO) complex can build up in mouse inducible nitric oxide synthase (iNOS) during NO synthesis from L-arginine. We investigated its formation kinetics, effect on catalytic activity, dependence on solution NO concentration, and effect on enzyme oxygen response (apparent KmO(2)). Heme-NO complex formation was biphasic and was linked kinetically to an inhibition of electron flux and catalysis in iNOS. Experiments that utilized a superoxide generating system to scavenge NO showed that the magnitude of heme-NO complex formation directly depended on the NO concentration achieved in the reaction solution. However, a minor portion of heme-NO complex (20%) still formed during NO synthesis even when solution NO was completely scavenged. Formation of the intrinsic heme-NO complex, and the heme-NO complex related to buildup of solution NO, increased the apparent KmO2 of iNOS by 10- and 4-fold, respectively. Together, the data show heme-NO complex buildup in iNOS is due to both intrinsic NO binding and to equilibrium binding of solution NO, with the latter predominating when NO reaches high nanomolar to low micromolar concentrations. This behavior distinguishes iNOS from the other NOS isoforms and indicates a more complex regulation is possible for its activity and oxygen response in biologic settings.