Two univalent electron transfers from putidaredoxin to bacterial cytochrome P450 at subzero temperature.
Two univalent electron transfers from putidaredoxin to bacterial cytochrome P450 at subzero temperature.
复制标题
在零下温度下从恶臭氧还蛋白到细菌细胞色素 P450 的两个单价电子转移。
DOI：
10.1021/bi00607a021
复制
发表时间：
1978
期刊：
影响因子：
2.9
通讯作者：
I. C. Gunsalus
中科院分区：
文献类型：
--
作者：
G. H. Hoa;E. Bégard;P. Debey;I. C. Gunsalus
一个完整的动态描述的多步骤多酶O2还原和羟基化序列仍然是一个挑战，值得多种方法。在零度以下的流体介质中进行的实验允许人们利用活化能的差异，并将整个过程分解为天然电子供体putidaredoxin对细胞色素P450的两个单价还原。我们首先考虑P450细胞色素-底物复合物的初始铁/亚铁还原。在-10至-40 ℃的温度范围内将化学还原的putidaredoxin与P450快速混合后分析动力学.随着初始putidaredoxin水平的增加，亚铁P450产率达到接近60%降低的明显平台。动力学分析表明，主要的，限速的，事件是形成一个双酶复合物，然后是一个可逆的分子间电子转移。还原速率直接取决于血红素P450偶联单加氧酶中1至25 µ单位氧还原中的putidaredoxin浓度，通过交替底物添加和单价还原的有序循环顺序已经有充分的文献记载（Estabrook埃塔尔，1968年; Gunsalus埃塔尔，1971，1974; Peterson等人，1973年）。模型系统对于阐明调节P450单加氧酶过程的动力学和热力学参数的重要性，通过光学和其他光学性质所暗示的催化中心的多样性、普遍性和相似性来强调。
A full dynamic description of the multistep multienzyme O2 reduction and hydroxylation sequence remains a challenge worthy of multiple approaches. Experimentation in fluid media at subzero temperatures allows one to exploit the differences in activation energy and to resolvethe overall process into thetwo univalent reductions of the cytochrome P450 by the natural electron donor, putidaredoxin. We consider first the initial ferric/ferrous reduction of a P450 cytochrome-substrate complex. The kinetics are analyzed after rapid mixing of chemically reduced putidaredoxin with the P450 over the temperature range—10 to—40 C. With in-creases in the initial putidaredoxin level, the ferrous P450 yield reaches an apparent plateau near 60% reduction. Analysis of the kinetics indicates the primary, and rate-limiting, event is the formationof a dienzyme complex followed by a reversible intermolecular electron transfer. The reduction rate depends directly on the putidaredoxin concentration in the 1 to 25 µDioxygen reduction inheme P450 coupled monoxygenases via an ordered cyclical sequence of alternate substrate addition and univalent reduction has been well documented (Estabrook etal., 1968; Gunsalus etal., 1971, 1974; Peterson et al., 1973). The importance of a model system to elucidate the kinetic and thermodynamic parameters that regulate the P450 monoxy-genase process is emphasized by the multiplicity, ubiquity, and similarities in the catalytic centers implied by optical and other