NITRIC-OXIDE COMPLEXES OF INDUCIBLE NITRIC-OXIDE SYNTHASE - SPECTRAL CHARACTERIZATION AND EFFECT ON CATALYTIC ACTIVITY
NITRIC-OXIDE COMPLEXES OF INDUCIBLE NITRIC-OXIDE SYNTHASE - SPECTRAL CHARACTERIZATION AND EFFECT ON CATALYTIC ACTIVITY
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DOI：
10.1021/bi00016a038
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发表时间：
1995-04-25
期刊：
影响因子：
2.9
通讯作者：
MARLETTA, MA
中科院分区：
文献类型：
--
作者：
HURSHMAN, AR;MARLETTA, MA
一氧化氮合酶（NOS）催化L-精氨酸氧化为瓜氨酸和一氧化氮（NO）。 NOS 是一种含有细胞色素 P-450 型血红素的血红素蛋白，已证明其参与催化作用。有人建议。 NO 能够与 NOS 的血红素紧密结合，并可能以这种方式调节酶活性。我们在这里报告了铁血红素和亚铁血红素亚硝酰复合物与小鼠巨噬细胞的诱导型 NOS 的形成。亚硝酰铁络合物的特征在于 443 nm 处的 Soret 峰以及 549 和 585 nm 处的 α/β 区域中的两个不同峰。亚硝酰亚铁络合物在 436 和 566 nm 处具有最大吸光度。在 NOS 转换的条件下观察到瞬态光谱中间体。该中间体似乎是亚硝酰亚铁络合物和亚铁络合物的混合物，并且在氧气存在下不稳定。 L-精氨酸的结合降低了 的亲和力。 NO 对于铁血红素但似乎不会降低 的亲和力。对于亚铁血红素而言，否。在 NOS 测定中添加氧合血红蛋白或高铁血红蛋白会导致酶活性增加近 2 倍。这一结果归因于两种形式的血红蛋白都能降低 的浓度。溶液中的 NO 与 一致。在测定条件下 NO 对 NOS 的抑制。我们的结果表明，NOS 亚硝酰复合物在某些条件下形成，但表明此类复合物与体内活性的相关性可能因其在有氧环境中的不稳定性而受到限制。
Nitric oxide synthase (NOS) catalyzes the oxidation of L-arginine to citrulline and nitric oxide (. NO). NOS is a hemoprotein containing a cytochrome P-450-type heme that has been shown to be involved in catalysis. It has been suggested that . NO is able to bind tightly to the heme of NOS and may in this way serve to regulate enzymatic activity. We report here the formation of both ferric and ferrous heme nitrosyl complexes with the inducible NOS from murine macrophages. The ferric nitrosyl complex is characterized by a Soret peak at 443 nm and two distinct peaks in the alpha/beta region at 549 and 585 nm. The ferrous nitrosyl complex has absorbance maxima at 436 and 566 nm. A transient spectral intermediate is observed under conditions of NOS turnover. This intermediate appears to be a mixture of ferric and ferrous nitrosyl complexes and is unstable in the presence of oxygen. Binding of L-arginine decreases the affinity of . NO for the ferric heme but does not appear to decrease the affinity of . NO for the ferrous heme. Addition of either oxyhemoglobin or methemoglobin to NOS assays results in a nearly 2-fold increase in enzymatic activity. This result is attributed to the ability of both forms of hemoglobin to decrease the concentration of . NO in solution and is consistent with . NO inhibition of NOS under assay conditions. Our results show that NOS nitrosyl complexes form under certain conditions but suggest that the relevance of such complexes to activity in vivo may be limited by their instability in an aerobic environment.