OXYGEN-UPTAKE AND ANTIOXIDANT RESPONSES OF THE FREE-LIVING DIPLOMONADHEXAMITA SP

The free-living anaerobic flagellate Hexamita sp. was observed to acti vely consume O-2 with a K-m O-2 of 13 mu M. Oxygen consumption increas ed linearly with O-2 tension up to a threshold level of 100 mu M, abov e which it was inhibited. Oxygen uptake was supported by a number of s ubstrates but probably not coupled to energy conservation as cytochrom es could not be detected spectrophotometrically. In addition, inhibito rs specific for respiratory chain components did not significantly aff ect O-2 uptake. Respiration was however, partially inhibited by flavop rotein and iron-sulfur protein inhibitors. NAD(P)H supported O-2 consu mption was measured in both particulate and soluble fractions; this ac tivity was partially inhibited by quinacrine. A chemosensory response was observed in cells exposed to air, however no response was observed in the presence of superoxide dismutase plus catalase. Catalase and n onspecific peroxidase activity could not be detected, but superoxide d ismutase activity was present. Superoxide dismutase was sensitive to N aN3 and H2O2 but not KCN, suggesting a Fe prosthetic group. Flow cytom etric analysis revealed that thiol levels in live cells were depleted in the presence of t-butyl H2O2. The observed NADPH-driven glutathione reductase activity is believed to recycle oxidized thiols in order to re-establish reduced thiol levels in the cell. The corresponding thio l cycling enzyme glutathione peroxidase could not be detected. The abi lity to withstand high O-2 tensions (100 mu M) would enable Hexamita t o spend short periods in a wider range of habitats. Prolonged exposure to O-2 tensions higher than 100 mu M leads to irreversible damage and cell death.