Nitric oxide binding to the heme of neuronal nitric-oxide synthase links its activity to changes in oxygen tension
Nitric oxide binding to the heme of neuronal nitric-oxide synthase links its activity to changes in oxygen tension
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DOI：
10.1074/jbc.271.51.32515
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发表时间：
1996-12-20
影响因子：
4.8
通讯作者：
Stuehr, DJ
中科院分区：
文献类型：
--
作者：
AbuSoud, HM;Rousseau, DL;Stuehr, DJ
神经元一氧化氮合酶（NOS-1）是一种血红素蛋白，可由L-精氨酸、O-2和NADPH产生NO和瓜氨酸。在催化过程中，大部分NOS-1结合自生NO并转化为亚铁-NO复合物，这导致其在稳态期间以其最大可能活性的一部分操作（Abu-Soud，H，M，，Wang，J，Rousseau，D，L，Fukuto，J，，M，L，J.，和Stuehr，D，J，（1995）J. Biol，Chem，270，22997-23006）。为了研究NO复合物的形成如何影响NOS-1的O-2响应，我测量了在存在和不存在L-精氨酸的情况下NO合成和NADPH氧化的速率与O-2浓度的关系。在不存在L-精氨酸的情况下，NOS-1催化简单的O-2还原，其血红素铁显示出典型的O-2亲和力（估计K-mO-2小于或等于40 μ M，饱和度接近100 μ M）。在L-精氨酸存在下，NO合成和NADPH氧化速率在更宽的范围内与O-2浓度成正比（估算的KmO 2类似于400 μ M，饱和度类似于800 μ M），表明亚铁-NO复合物的形成改变了NOS-1的O-2响应，停流实验表明，在100 - 700 μ M之间，亚铁-NO络合物的形成速率与O-2浓度相对无关，而复合物的分解速率与O-2浓度成正比。我们的结论是铁-NO复合物的O-2敏感性控制了NOS-1的O-2响应，从而在稳态期间控制了其活性，这使得NOS-1能够在整个生理范围内将其NO合成速率与O-2浓度耦合。
Neuronal nitric-oxide synthase (NOS-1) is a hemeprotein that generates NO and citrulline from L-arginine, O-2, and NADPH. During catalysis, a majority of NOS-1 binds self-generated NO and converts to a ferrous-NO complex, which causes it to operate at a fraction of its maximum possible activity during the steady state (Abu-Soud, H, M,, Wang, J,, Rousseau, D, L,, Fukuto, J,, Ignarro, L, J., and Stuehr, D, J, (1995) J. Biol, Chem, 270, 22997-23006), To examine how NO complex formation affects the O-2 response of NOS-1, me measured rates of NO synthesis and NADPH oxidation versus O-2 concentration in the presence and absence of L-arginine, In the absence of L-arginine, NOS-1 catalyzed simple O-2 reduction, and its heme iron displayed a typical affinity for O-2 (estimated K-m O-2 less than or equal to 40 mu M, saturation at similar to 100 mu M). In the presence of L-arginine, the rates of NO synthesis and NADPH oxidation were proportional to the O-2 concentration over a much broader range (estimated KmO2 similar to 400 mu M, saturation at similar to 800 mu M), indicating that ferrous-NO complex formation altered the O-2 response of NOS-1, Stopped-flow experiments revealed that the rate of ferrous-NO complex formation was relatively independent of the O-2 concentration between 100 and 700 mu M, while the rate of complex breakdown was directly proportional to O-2 concentration. We conclude that the O-2 sensitivity of the ferrous-NO complex governs the O-2 response of NOS-1 and thus its activity during the steady state, This enables NOS-1 to couple its rate of NO synthesis to the O-2 concentration throughout the physiologic range.