Serra M C, Bellavite P, Davoli A, Bannister J V, Rossi F
Biochim Biophys Acta. 1984 Jul 17;788(1):138-46. doi: 10.1016/0167-4838(84)90306-6.
NADPH-dependent O2- forming activity was extracted with deoxycholate from subcellular particles of guinea-pig neutrophils following stimulation with phorbol myristate acetate. The solubilized enzyme was purified by chromatography on Ultrogel AcA22, by isopycnic glycerol density gradient centrifugation and by treatment with 0.4 M NaCl. This procedure yielded a high-molecular-weight complex containing phospholipids, cytochrome b-245 and NADPH oxidase activity. Cytochrome b was found to be purified to the same extent as NADPH oxidase activity. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the various purification fractions showed a progressive enrichment of a band whose molecular weight is 3.2 X 10(4). The enrichment of this protein band paralleled those of NADPH oxidase activity and of cytochrome b, indicating that it is a component of the oxidase system. The possibility that this band corresponds to either cytochrome b or a flavoprotein/cytochrome b complex is considered.
在用佛波醇肉豆蔻酸酯乙酸盐刺激后，从豚鼠中性粒细胞的亚细胞颗粒中用脱氧胆酸盐提取依赖烟酰胺腺嘌呤二核苷酸磷酸（NADPH）的产氧活性。通过在Ultrogel AcA22上进行色谱分离、等密度甘油密度梯度离心以及用0.4M氯化钠处理，对溶解的酶进行纯化。该过程产生了一种含有磷脂、细胞色素b - 245和NADPH氧化酶活性的高分子量复合物。发现细胞色素b的纯化程度与NADPH氧化酶活性相同。对各种纯化级分进行十二烷基硫酸钠聚丙烯酰胺凝胶电泳显示，一条分子量为3.2×10⁴的条带逐渐富集。该蛋白条带的富集与NADPH氧化酶活性和细胞色素b的富集平行，表明它是氧化酶系统的一个组分。考虑了这条带对应于细胞色素b或黄素蛋白/细胞色素b复合物的可能性。
