Mayer B, Klatt P, Werner E R, Schmidt K
Institut für Pharmakologie und Toxikologie, Karl-Franzens-Universität Graz, Austria.
J Biol Chem. 1995 Jan 13;270(2):655-9. doi: 10.1074/jbc.270.2.655.
A Clark-type nitric oxide-sensitive electrode was used for electrochemical determination of NO oxidation kinetics. Reaction with molecular oxygen followed second-order rate law with respect to NO with an overall rate constant of 9.2 +/- 0.33 x 10(6) M-2 s-1. Tetrahydrobiopterin, an essential cofactor of NO synthases, was found to induce rapid oxidation of NO in a 1:1 stoichiometry. The reaction required the presence of oxygen, was zero order with respect to NO and first order with respect to tetrahydrobiopterin, completely blocked by 5,000 units/ml superoxide dismutase, and mimicked by a superoxide-generating system. Purified brain NO synthase produced no detectable NO unless high amounts of superoxide dismutase were present. NO synthase-catalyzed citrulline formation was inhibited by superoxide dismutase (5,000 units/ml) in an oxyhemoglobin-sensitive manner, indicating that NO induces feedback inhibition of NO synthase. NO-stimulated soluble guanylyl cyclase was inhibited by tetrahydrobiopterin at half-maximally active concentrations of 2 microM. The present data suggest that NO is inactivated to peroxynitrite by superoxide generated in the course of tetrahydrobiopterin autoxidation.
使用Clark型一氧化氮敏感电极对一氧化氮氧化动力学进行电化学测定。一氧化氮与分子氧的反应遵循关于一氧化氮的二级速率定律，总速率常数为9.2±0.33×10(6) M-2 s-1。四氢生物蝶呤是一氧化氮合酶的必需辅因子，被发现能以1:1的化学计量比诱导一氧化氮快速氧化。该反应需要氧气的存在，对一氧化氮为零级反应，对四氢生物蝶呤为一级反应，完全被5000单位/毫升的超氧化物歧化酶阻断，并被超氧化物生成系统模拟。纯化的脑一氧化氮合酶除非存在大量超氧化物歧化酶，否则不会产生可检测到的一氧化氮。一氧化氮合酶催化的瓜氨酸形成受到超氧化物歧化酶（5000单位/毫升）以氧合血红蛋白敏感的方式抑制，表明一氧化氮诱导对一氧化氮合酶的反馈抑制。在2微摩尔的半最大活性浓度下，四氢生物蝶呤抑制一氧化氮刺激的可溶性鸟苷酸环化酶。目前的数据表明，一氧化氮在四氢生物蝶呤自氧化过程中产生的超氧化物作用下被灭活为过氧亚硝酸盐。
