Mason Maria G, Nicholls Peter, Wilson Michael T, Cooper Christopher E
Department of Biological Sciences, University of Essex, Wivenhoe Park, Colchester CO4 3SQ, United Kingdom.
Proc Natl Acad Sci U S A. 2006 Jan 17;103(3):708-13. doi: 10.1073/pnas.0506562103. Epub 2006 Jan 9.
NO reversibly inhibits mitochondrial respiration via binding to cytochrome c oxidase (CCO). This inhibition has been proposed to be a physiological control mechanism and/or to contribute to pathophysiology. Oxygen reacts with CCO at a heme iron:copper binuclear center (a(3)/Cu(B)). Reports have variously suggested that during inhibition NO can interact with the binuclear center containing zero (fully oxidized), one (singly reduced), and two (fully reduced) additional electrons. It has also been suggested that two NO molecules can interact with the enzyme simultaneously. We used steady-state and kinetic modeling techniques to reevaluate NO inhibition of CCO. At high flux and low oxygen tensions NO interacts predominantly with the fully reduced (ferrous/cuprous) center in competition with oxygen. However, as the oxygen tension is raised (or the consumption rate is decreased) the reaction with the oxidized enzyme becomes increasingly important. There is no requirement for NO to bind to the singly reduced binuclear center. NO interacts with either ferrous heme iron or oxidized copper, but not both simultaneously. The affinity (K(D)) of NO for the oxygen-binding ferrous heme site is 0.2 nM. The noncompetitive interaction with oxidized copper results in oxidation of NO to nitrite and behaves kinetically as if it had an apparent affinity of 28 nM; at low levels of NO, significant binding to copper can occur without appreciable enzyme inhibition. The combination of competitive (heme) and noncompetitive (copper) modes of binding enables NO to interact with mitochondria across the full in vivo dynamic range of oxygen tension and consumption rates.
一氧化氮（NO）通过与细胞色素c氧化酶（CCO）结合可逆地抑制线粒体呼吸。这种抑制作用被认为是一种生理控制机制和/或对病理生理学有影响。氧气在血红素铁：铜双核中心（a(3)/Cu(B)）与CCO发生反应。有报道指出，在抑制过程中，NO可以与含有零个（完全氧化）、一个（单电子还原）和两个（完全还原）额外电子的双核中心相互作用。也有人提出两个NO分子可以同时与该酶相互作用。我们使用稳态和动力学建模技术重新评估了NO对CCO的抑制作用。在高通量和低氧张力下，NO主要与完全还原的（亚铁/亚铜）中心相互作用，与氧气竞争。然而，随着氧张力升高（或消耗速率降低），与氧化态酶的反应变得越来越重要。NO不需要与单电子还原的双核中心结合。NO与亚铁血红素铁或氧化态铜相互作用，但不会同时与两者相互作用。NO对氧结合亚铁血红素位点的亲和力（K(D)）为0.2 nM。与氧化态铜的非竞争性相互作用导致NO氧化为亚硝酸盐，其动力学行为就好像它的表观亲和力为28 nM；在低水平的NO下，与铜的显著结合可以在没有明显酶抑制的情况下发生。竞争性（血红素）和非竞争性（铜）结合模式的结合使NO能够在体内氧气张力和消耗速率的整个动态范围内与线粒体相互作用。
