Morel F, Vignais P V
Biochim Biophys Acta. 1984 Feb 27;764(2):213-25. doi: 10.1016/0005-2728(84)90030-6.
The spectral properties of a particulate fraction of human polymorphonuclear neutrophils capable of oxidizing NADPH were studied before and after depletion of myeloperoxidase by KCl treatment. Difference spectra (dithionite reduced minus oxidized) at 77 K of non-extracted particles showed peaks of a b-type cytochrome at 556, 527 and 425 nm and of myeloperoxidase at 636 and 474 nm. Extraction of myeloperoxidase led to a 4-5-fold increase in the size of the cytochrome b peaks. In non-extracted particles, the CO-reduced spectra at 77 K revealed a typical CO-reduced myeloperoxidase complex with new peaks at 625-630 and 462 nm, and a limited shift of the Soret band of reduced cytochrome b from 425 to 424-423 nm. The same shift was observed for cytochrome b in extracted particles. Photoirradiation of the CO-dithionite-reduced particles resulted in a back shift of the CO-reduced peaks to their original positions in the reduced spectrum. Concomitantly, the size of the peaks both for myeloperoxidase and cytochrome b was increased, indicating photoreduction. Cytochrome b and myeloperoxidase in neutrophil particles were poorly reduced by NADPH; reduction occurred upon photoirradiation. FAD and FMN added to particles in the presence of NADPH were photoreduced concomitantly with cytochrome b. Addition of phorbol myristate acetate to intact neutrophils in the presence of glucose resulted in CO- and cyanide-insensitive respiration, accumulation of O-2, and also in reduction of cytochrome b. The lag required to reach the steady-state production of O-2 was equal to the lag required for cytochrome b to reach a plateau of reduction. The data are consistent with the idea that cytochrome b in neutrophils might belong to a branched pathway that is not rate-limiting in the cyanide-resistant respiration of the neutrophils.
在通过氯化钾处理使髓过氧化物酶耗竭之前和之后，研究了能够氧化NADPH的人多形核中性粒细胞颗粒部分的光谱特性。未提取颗粒在77K下的差示光谱（连二亚硫酸盐还原减去氧化）显示，b型细胞色素在556、527和425nm处有峰，髓过氧化物酶在636和474nm处有峰。髓过氧化物酶的提取导致细胞色素b峰的大小增加了4至5倍。在未提取颗粒中，77K下的一氧化碳还原光谱显示出典型的一氧化碳还原髓过氧化物酶复合物，在625 - 630和462nm处有新峰，并且还原型细胞色素b的Soret带从425nm有限地移动到424 - 423nm。在提取颗粒中的细胞色素b也观察到相同的移动。一氧化碳 - 连二亚硫酸盐还原颗粒的光照射导致一氧化碳还原峰在还原光谱中回到其原始位置。同时，髓过氧化物酶和细胞色素b的峰大小都增加，表明光还原。中性粒细胞颗粒中的细胞色素b和髓过氧化物酶被NADPH还原的程度很低；光照射时发生还原。在NADPH存在下添加到颗粒中的FAD和FMN与细胞色素b同时被光还原。在葡萄糖存在下向完整的中性粒细胞中添加佛波醇肉豆蔻酸酯导致对一氧化碳和氰化物不敏感的呼吸、O₂的积累，并且还导致细胞色素b的还原。达到O₂稳态产生所需的延迟等于细胞色素b达到还原平台所需的延迟。这些数据与中性粒细胞中的细胞色素b可能属于在中性粒细胞抗氰呼吸中不限制速率的分支途径的观点一致。
