The intra-mitochondrial localization of flavoproteins previously assigned to the respiratory chain.

Abstract

1 Flavoproteins of intact and fractionated mitochondria have been studied by means of spectrophotometric and fluorimetric techniques. 2 The high and low redox potential fluorescent flavoproteins of rat liver mitochondria previously assigned to the respiratory chain are located in the soluble fraction obtained on ultrasonic disintegration. 3 In intact mitochondria, the low potential species has a redox potential of −279 mV at pH 7.2 and 25° measured by fluorescence or absorbance, and undergoes a 2 equivalent reduction. After ultrasonic disruption of mitochondria, its redox potential is -294 m V at pH 7.2 and 25°. 4 In submitochondrial particles, low potential flavoprotein reducible by NADH in the presence of rotenone is not fluorescent, appears to have a redox potential more positive than −209 mV and is attributed mainly to cytochrome b5 reductase and cytochrome b5. Mersalyl can be used to selectively inhibit cytochrome b5 reductase, and it has been possible to estimate the concentration of the low redox potential flavoprotein free from cytochrome b5 interference. 5 The high redox potential flurescent flavoprotein is recovered in the soluble fraction after ultrasonic disruption of rat liver mitochondria and can be reduced by succinate in the presence of antimycin A inhibited particulate fraction. This fluorescent species is also found in rat kidney mitochondria but not in blowfly mitochondria. 6 In submitochondrial particles, a non-fluorescent high redox potential species is found which may be flavoprotein or non-heme iron. This species has a redox potential of + 30 m V at pH 7.2 and 25° and undergoes 1 equivalent reduction. 7 Flavoprotein fluorescence measurements are subject to significant errors caused by absorption of the exciting light, and conseuently do not always measure flavoproteins free from interference by other respiratory carriers, e. q. cytochromes and non-heme iron proteins.