PARTIAL-PURIFICATION OF THE CYANIDE-RESISTANT ALTERNATIVE OXIDASE OF SKUNK CABBAGE (SYMPLOCARPUS-FOETIDUS) MITOCHONDRIA
PARTIAL-PURIFICATION OF THE CYANIDE-RESISTANT ALTERNATIVE OXIDASE OF SKUNK CABBAGE (SYMPLOCARPUS-FOETIDUS) MITOCHONDRIA
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DOI：
10.1104/pp.101.1.113
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发表时间：
1993-01-01
期刊：
影响因子：
7.4
通讯作者：
SIEDOW, JN
中科院分区：
文献类型：
--
作者：
BERTHOLD, DA;SIEDOW, JN
从臭鼬甘蓝（Symplocarpus foetidus L）肉穗花序线粒体中部分纯化了抗氰化物的交替氧化酶。臭鼬甘蓝线粒体溶解在N-双-（3-D-葡糖酰胺丙基）脱氧胆酰胺和交替氧化酶纯化使用一批DEAE-纤维素处理，然后沉淀与Extracti-Gel和Sephadex G-200上的色谱。汇集和浓缩的最活跃的馏分从凝胶过滤柱，一个20- 30倍的纯化的替代氧化酶获得，没有证据污染细胞色素c氧化酶（复合物IV）或细胞色素c还原酶（复合物III）。部分纯化的氧化酶的聚丙烯酰胺凝胶电泳显示在36和29 kD的主要多肽，这两个反应与单克隆抗体提出对Sauromatum guttatum交替氧化酶。纯化的氧化酶级分在可见光谱区没有吸光度，并且加入硼氢化钠在紫外区没有吸光度变化。纯化的交替氧化酶催化的四电子还原的氧到水的情况下，柠檬酸盐，但催化的一个明显的两电子还原的氧到过氧化氢的存在下，0.7 M柠檬酸盐。
A partial purification of the cyanide-resistant, alternative oxidase from skunk cabbage (Symplocarpus foetidus L) spadix mitochondria is described. Skunk cabbage mitochondria were solubilized in NN-bis-(3-D-glucon-amido-propyl)deoxycholamide and the alternative oxidase was purified using a batch DEAE-cellulose treatment, followed by precipitation with Extracti-Gel and chromatography on Sephadex G-200. Following pooling and concentrating of the most active fractions from the gel filtration column, a 20- to 30-fold purification of the alternative oxidase was obtained, with no evidence of contamination by cytochrome c oxidase (complex IV) or cytochrome c reductase (complex III). Polyacrylamide gel electrophoresis of the partially purified oxidase showed major polypeptides at 36 and 29 kD, both of which react with monoclonal antibodies raised against the Sauromatum guttatum alternative oxidase. The purified oxidase fraction showed no absorbance in the visible spectral region, and addition of sodium borohydride induced no absorbance changes in the ultraviolet region. The purified alternative oxidase catalyzed the four-electron reduction of oxygen to water in the absence of citrate, but catalyzed an apparent two-electron reduction of oxygen to hydrogen peroxide in the presence of 0.7 m citrate.