Wang J, Rousseau D L, Abu-Soud H M, Stuehr D J
AT&T Bell Laboratories, Murray Hill, NJ 07974.
Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10512-6. doi: 10.1073/pnas.91.22.10512.
A current question in nitric oxide (NO) biology is whether NO can act as a feedback inhibitor of NO synthase (NOS). We have approached this problem by examining the interaction of NO with neuronal NOS by optical absorption and resonance Raman scattering spectroscopies. Under an inert atmosphere NO coordinated to the heme iron in both the oxidized and reduced forms of NOS. The Soret and visible optical absorption transitions are detected at 436 and at 567 nm, respectively, in the Fe(2+)-NO heme complex and at 440 nm and at 549 and 580 nm, respectively, in the Fe(3+)-NO heme complex. In the resonance Raman spectrum of the ferrous complex the Fe-NO stretching mode is located at 549 cm-1 in the presence of L-arginine and at 536 cm-1 in the absence of L-arginine, whereas in the ferric enzyme the mode is located at 540 cm-1 (in the absence of L-arginine). The interaction between bound L-arginine and the NO indicates that L-arginine binds directly over the heme just as do the substrates in cytochrome P-450s. In the absence of L-arginine, NO readily oxidized the ferrous heme iron. The oxidation was prevented by the presence of bound L-arginine and enabled NOS to form a stable ferrous NO complex. Under oxygen-limited conditions, NO generated by neuronal NOS coordinated to its heme iron and formed a spectrally detectable ferrous-NO complex. Taken together, our results show that NO can bind to both ferric and ferrous NOS and may inhibit NO synthesis through its binding to the heme iron during catalysis.
一氧化氮（NO）生物学领域当前的一个问题是，NO是否能作为一氧化氮合酶（NOS）的反馈抑制剂。我们通过光学吸收光谱和共振拉曼散射光谱研究NO与神经元型NOS的相互作用来解决这个问题。在惰性气氛下，NO与氧化态和还原态的NOS中的血红素铁配位。在Fe(2+)-NO血红素复合物中，Soret和可见光吸收跃迁分别在436和567 nm处检测到，而在Fe(3+)-NO血红素复合物中分别在440 nm以及549和580 nm处检测到。在亚铁复合物的共振拉曼光谱中，Fe-NO伸缩模式在存在L-精氨酸时位于549 cm-1，在不存在L-精氨酸时位于536 cm-1，而在高铁酶中该模式位于540 cm-1（不存在L-精氨酸时）。结合的L-精氨酸与NO之间的相互作用表明，L-精氨酸就像细胞色素P-450中的底物一样直接结合在血红素上方。在不存在L-精氨酸的情况下，NO很容易氧化亚铁血红素铁。结合的L-精氨酸的存在可防止这种氧化，并使NOS形成稳定的亚铁NO复合物。在氧气受限的条件下，神经元型NOS产生的NO与其血红素铁配位并形成可通过光谱检测的亚铁-NO复合物。综上所述，我们的结果表明，NO可以与高铁和亚铁NOS结合，并且可能在催化过程中通过与血红素铁结合来抑制NO的合成。
