Class I heme peroxidases: Characterization of soybean ascorbate peroxidase
Class I heme peroxidases: Characterization of soybean ascorbate peroxidase
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DOI：
10.1006/abbi.1998.0941
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发表时间：
1998-12-15
影响因子：
3.9
通讯作者：
Raven, EL
中科院分区：
文献类型：
--
作者：
Jones, DK;Dalton, DA;Raven, EL
高效表达系统[D. A.道尔顿等.生物化学.生物物理. 328，1-8，1996）首次用于大豆根瘤抗坏血酸过氧化物酶（APX）以产生足够大量的酶用于详细的生物物理分析。重组APX已通过电子吸收，EPR，NMR和圆二色谱法表征，并通过电化学，电子，EPR和NMR光谱与298 R处酶的高自旋铁静止状态一致。低温EPR（7 K）和电子吸收（77 K）实验表明，在这些温度下形成的低自旋血红素衍生物。通过光谱电化学测定，Ee（III）/Fe（II）氧化还原电对的中点还原电位为-159 +/- 2 mV vs SHE（pH 7.0，25.0摄氏度，μ =0.10 M）。豌豆和大豆APX的圆二色谱非常相似，表明这两种酶具有共同的结构特征。大豆APX的熔化温度，通过圆二色光谱监测，是49摄氏度，这些结果代表了第一个详细的光谱和电化学分析大豆抗坏血酸过氧化物酶，并在更广泛的范围内讨论其他I类过氧化物酶。(C)北京：科学出版社.
An efficient expression system [D. A. Dalton ct al. Arch. Biochem. Biophys. 328, 1-8, 1996) for soybean nodule ascorbate peroxidase (APX) has, for the first time, been used to generate enzyme in large enough quantities for detailed biophysical analysis. The recombinant APX has been characterized by electronic absorption, EPR, NMR and circular dichroism spectroscopies, and by electrochemistry, Electronic, EPR, and NMR spectra are consistent with a high-spin ferric resting state for the enzyme at 298 R. Low-temperature EPR (7 K) and electronic absorption (77 K) experiments indicate formation of a low-spin heme derivative at these temperatures. The midpoint reduction potential for the Ee(III)/Fe(II) redox couple, determined by spectroelectrochemistry, is -159 +/- 2 mV vs SHE (pH 7.0, 25.0 degrees C, mu =0.10 M). Circular dichroism spectra of pea and soybean APXs are very similar, indicating common structural features for the two enzymes. The melting temperature of soybean APX, as monitored by circular dichroism spectroscopy, is 49 degrees C, These results represent the first detailed spectroscopic and electrochemical analysis of soybean ascorbate peroxidase and are discussed in the broader context of other class I peroxidases. (C) 1998 Academic Press.