Ellis J A, Cross A R, Jones O T
Department of Biochemistry, Medical School, University of Bristol, U.K.
Biochem J. 1989 Sep 1;262(2):575-9. doi: 10.1042/bj2620575.
A superoxide-generating NADPH oxidase was solubilized from phorbol 12-myristate 13-acetate-activated human neutrophils with a mixture of sodium deoxycholate (0.125%, w/v) and Lubrol-PX (0.125%, v/v). The solubilized preparation contained FAD (577 pmol/mg of protein) and cytochrome b-245 (479 pmol/mg of protein) and produced 11.61 mol of O2-./s per mol of cytochrome b (340 nmol of O2-./min per mg of protein). On addition of NADPH, the cytochrome b-245 was reduced by 7.9% and the FAD by 38% in the aerobic steady state; NADH addition caused little steady-state reduction of cytochrome b and FAD. In this preparation, and several others, the measured rate of O2-. production correlated with the turnover of cytochrome b calculated from the extent of cytochrome b-245 reduction under aerobic conditions. Addition of diphenyleneiodonium abolished the reduction of both the FAD and cytochrome b-245 components and inhibited O2-. production. The haem ligand imidazole inhibited O2-. generation and cytochrome b reduction while permitting FAD reduction. These results support the suggestion that the human neutrophil NADPH oxidase has the electron-transport sequence: NADPH----FAD----cytochrome b-245----O2.
用脱氧胆酸钠（0.125%，w/v）和Lubrol - PX（0.125%，v/v）的混合物从佛波酯12 - 肉豆蔻酸酯13 - 乙酸酯激活的人中性粒细胞中溶解出一种产生超氧化物的NADPH氧化酶。溶解后的制剂含有FAD（577 pmol/mg蛋白质）和细胞色素b - 245（479 pmol/mg蛋白质），每摩尔细胞色素b每秒产生11.61摩尔O₂⁻（每毫克蛋白质每分钟产生340 nmol O₂⁻）。在有氧稳态下加入NADPH后，细胞色素b - 245减少7.9%，FAD减少38%；加入NADH对细胞色素b和FAD的稳态减少作用很小。在该制剂以及其他几种制剂中，测得的O₂⁻产生速率与根据有氧条件下细胞色素b - 245的还原程度计算出的细胞色素b周转率相关。加入二亚苯基碘鎓可消除FAD和细胞色素b - 245成分的还原，并抑制O₂⁻的产生。血红素配体咪唑抑制O₂⁻的产生和细胞色素b的还原，同时允许FAD还原。这些结果支持了人中性粒细胞NADPH氧化酶具有电子传递序列：NADPH→FAD→细胞色素b - 245→O₂的观点。
