ISOLATION, CHARACTERIZATION AND FUNCTION OF THE 2 CYTOCHROMES-C OF THE YEAST CANDIDA-PARAPSILOSIS

Candida parapsilosis is a strictly aerobic yeast which possesses two r espiratory chains with a peculiar organisation, different from that of plant mitochondria. Besides the classical electron transport pathway, mitochondria of C. parapsilosis develops an alternative pathway, whic h does not branch off at the ubiquinone level, but merges at the compl ex IV level. Two pools of cytochromes c were distinguished by their sp ectrometric and potentiometric properties: (i) sequential cytochrome c reduction was promoted by two substrates, PMS (E(m) = 70 mV) and TMPD (E(m) = 280 mV). TMPD promoted the reduction of a cytochrome c with m axima at 551.9 and 417.3 nm for the alpha and the Soret bands, respect ively, whereas cytochrome c reducible by PMS exhibited maxima at 549.7 and 419.9 nm: (ii) two midpoint redox potentials were resolved at 180 mV and 280 mV, respectively. The two cytochromes c were copurified by ion-exchange chromatography on Amberlite; after this step, the two cy tochromes c can always be differentiated by TMPD and PMS, these reduct ants promoting different absorption bands. The two cytochromes c were separated by reverse-phase HPLC; this last purification step resolved two proteins with the same relative molecular mass of 13600 but a diff erent amino-acid composition. Comparison of N-terminal sequences revea led differences between the two proteins. It was hypothesized that one cytochrome c is implicated in the functioning of the main chain and t he other in that of the secondary pathway.