A QUANTITATIVE TEST FOR SUPEROXIDE RADICALS PRODUCED IN BIOLOGICAL-SYSTEMS
A QUANTITATIVE TEST FOR SUPEROXIDE RADICALS PRODUCED IN BIOLOGICAL-SYSTEMS
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DOI：
10.1042/bj2030551
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发表时间：
1982-01-01
影响因子：
4.1
通讯作者：
ESTABROOK, RW
中科院分区：
文献类型：
--
作者：
KUTHAN, H;ULLRICH, V;ESTABROOK, RW
报道了一种适用于检测肝微粒体中超氧阴离子自由基的部分琥珀酰化细胞色素c的制备和性质。通过马心细胞色素c的45%的伯氨基的琥珀酰化，与天然细胞色素c相比，对溶解的NADPH-细胞色素P-450还原酶的活性降低了99%。细胞色素b5和细胞色素c氧化酶的能力，以减少琥珀酰化的铁细胞色素c和氧化琥珀酰化的铁细胞色素c，分别下降到类似的程度。然而，双分子速率常数为部分琥珀酰化的铁细胞色素c的O2-的还原。估计为O2-反应值的1/10。与天然铁细胞色素c在pH 7.7下反应。在此基础上，对O2-.由补充NADPH的[大鼠]肝微粒体产生。在各种有限浓度的细胞色素c衍生物确定的琥珀酰化铁细胞色素c还原的初始速率可以外推，以获得真正的O2-速率。在一个均匀的系统中生成。本文讨论了O_2 ~-定量测定中存在的问题。在生物膜，如微粒体中产生的，进行了讨论。
The preparation and properties of a partially succinoylated cytochrome c, suited for the detection of superoxide anion radicals in liver microsomes, is reported. By succinoylation of 45% of the primary amino groups of horse heart cytochrome c the activity towards solubilized NADPH-cytochrome P-450 reductase was diminished by 99% compared with native cytochrome c. The capacities of cytochrome b5 and cytochrome c oxidase to reduce the succinoylated ferricytochrome c and oxidize succinoylated ferrocytochrome c, respectively, were decreased to a similar extent. However, the bimolecular rate constant for the reduction of the partially succinoylated ferricytochrome c by O2-. was estimated to be 1/10 of the value for the reaction of O2-. with native ferricytochrome c at pH 7.7. On this basis the quantification of O2-. generated by NADPH-supplemented [rat] liver microsomes became possible. The initial rates of succinoylated ferricytochrome c reduction determined at various finite concentrations of the cytochrome c derivative can be extrapolated to obtain true rates of O2-. generation in a homogeneous system. The problems encountered in the quantitative determination of O2-. produced in biological membranes, e.g. microsomes, are discussed.