Wohlrab H, Greaney J
Biochim Biophys Acta. 1978 Sep 7;503(3):425-36. doi: 10.1016/0005-2728(78)90142-1.
Mitochondria have been prepared from the flight muscles of mature blow-flies (Sarcophaga bullata). Phosphate transport by these mitochondria, determined by rates of passive swelling in ammonium phosphate, is sensitive to inhibition by N-ethylmaleimide. 20 nmol of N-ethylmaleimide/nmol cytochrome A inhibit the swelling by 90%. When the mitochondria are inhibited by N-[3H]ethylmaleimide, then solubilized in dodecyl sulfate/mercaptoethanol at 100 degrees C and then electrophoresed on dodecyl sulfate-polyacrylamide gels, many labeled protein bands can be detected, including a large labeled peak that has the same mobility as the tracking dye, bromophenol blue. Sonic submitochondrial particles that are prepared from the N-[3H]ethylmaleimide-labeled mitochondria, solubilized, and electrophoresed on dodecyl sulfate-polyacrylamide gels, possess only seven major labeled protein bands with no radioactive peak at the tracking dye. These labeled proteins have molecular weights of 71, 68, 64, 45, 32, 30, and approx. 10 . 10(3). The nmol N-[3H]ethylmaleimide bound to each of these proteins per nmol cytochrome A are 0.15, 0.19, 0.35, 0.45, 0.87, 0.10, and 0.17, respectively, when the mitochondria are inhibited with 21.5 mol N-[3H]ethylmaleimide/mol cytochrome a at 10 micron cytochrome A. Coty and Pedersen (1975) J. Biol. Chem. 250, 3515-3521) sensitized rat liver mitochondria to N-[3H]ethylmaleimide and identified five labeled proteins. Only the labeled 32 . 10(3) dalton and the 45 . 10(3) dalton proteins are common to both systems.
已从成熟的麻蝇（肉蝇）飞行肌肉中制备出线粒体。这些线粒体的磷酸盐转运（通过在磷酸铵中被动肿胀的速率来测定）对N - 乙基马来酰亚胺的抑制敏感。每nmol细胞色素A有20 nmol的N - 乙基马来酰亚胺可抑制90%的肿胀。当线粒体被N - [³H]乙基马来酰亚胺抑制后，在100℃下用十二烷基硫酸钠/巯基乙醇溶解，然后在十二烷基硫酸钠 - 聚丙烯酰胺凝胶上进行电泳，可检测到许多标记蛋白条带，包括一个与示踪染料溴酚蓝迁移率相同的大的标记峰。由N - [³H]乙基马来酰亚胺标记的线粒体制备的超声亚线粒体颗粒，溶解后在十二烷基硫酸钠 - 聚丙烯酰胺凝胶上电泳，仅具有七条主要的标记蛋白条带，在示踪染料处没有放射性峰。这些标记蛋白的分子量分别为71、68、64、45、32、30和约10×10³。当线粒体在10μM细胞色素A的条件下用21.5 mol N - [³H]乙基马来酰亚胺/mol细胞色素a抑制时，每nmol细胞色素A与这些蛋白中每一种结合的N - [³H]乙基马来酰亚胺的nmol数分别为0.15、0.19、0.35、0.45、0.87、0.10和0.17。科蒂和佩德森（1975年，《生物化学杂志》250卷，3515 - 3521页）使大鼠肝线粒体对N - [³H]乙基马来酰亚胺敏感，并鉴定出五种标记蛋白。两个系统中仅有的共同标记蛋白是32×10³道尔顿和45×10³道尔顿的蛋白。
