Respiration of Trichomonas vaginalis. Components detected by electron paramagnetic resonance spectroscopy

Abstract

1 Electron paramagnetic resonance (EPR) spectrometry was used to characterise the iron-sulphur clusters present in the human parasite, Trichomonas vaginalis, an aerotolerant anaerobic protozoan which lacks mitochondria. 2 From observations of whole cells and subcellular fractions, the majority of the EPR signals are derived from one of the two main terminal respiration sites, the hydrogenosome; fractionation of hydrogenosomes revealed that a large proportion of these signals were associated with the membrane of these organelles. 3 Resolution into eight species was achieved by varying the degree of reduction and the temperature of the samples. 4 One component, a [2Fe–2S] ferredoxin reducible by pyruvate: ferredoxin oxidoreductase, corresponds to that previously purified, and predominates in spectra scanned at temperatures higher than 45 K. This species (mid-point redox potential – 300 ± 20 mV) was detected both in a hydrogenosomal membrane fraction and in the matrix fraction derived from broken organelles. 5 Another iron-sulphur species (mid-point potential – 270 mV) was detectable in both membrane and matrix at temperatures lower than 30 K. 6 Four other reduced species were confined to the hydrogenosomal membrane and a fifth was not detected after subfractionation, although it appeared in intact organelles. This species is assumed to be highly unstable. 7 Free-radical signals in hydrogenosome-enriched fractions probably arise from flavoprotein semiquinones; no other signals were obtained from the cytosolic fractions incubated with NADH. 8 A signal in the oxidised state was observed in the hydrogenosomal membrane. 9 Possible identities of hydrogenosomal iron-sulphur clusters are discussed in relation to previously established enzyme activities.