Characterization of the DNA gyrase from the thermoacidophilic archaeon Thermoplasma acidophilum.

Thermoplasma acidophilum is sensitive to the antibiotic drug novobiocin, which inhibits DNA gyrase. We characterized DNA gyrases from T. acidophilum strains in vitro. The DNA gyrase from a novobiocin-resistant strain and an engineered mutant were less sensitive to novobiocin. The novobiocin-resistant gyrase genes might serve as T. acidophilum genetic markers.

Effects of a squalene epoxidase inhibitor, terbinafine, on ether lipid biosyntheses in a thermoacidophilic archaeon, Thermoplasma acidophilum.

The archaeal plasma membrane consists mainly of diether lipids and tetraether lipids instead of the usual ester lipids found in other organisms. Although a molecule of tetraether lipid is thought to be synthesized from two molecules of diether lipids, there is no direct information about the biosynthetic pathway(s) or intermediates of tetraether lipid biosynthesis. In this study, we examined the effects of the fungal squalene epoxidase inhibitor terbinafine on the growth and ether lipid biosyntheses in the thermoacidophilic archaeon Thermoplasma acidophilum. Terbinafine was found to inhibit the growth of T. acidophilum in a concentration-dependent manner. When growing T. acidophilum cells were pulse-labeled with [2-(14)C]mevalonic acid in the presence of terbinafine, incorporation of radioactivity into the tetraether lipid fraction was strongly suppressed, while accumulation of radioactivity was noted at the position corresponding to diether lipids, depending on the concentration of terbinafine. After the cells were washed with fresh medium and incubated further without the radiolabeled substrate and the inhibitor, the accumulated radioactivity in the diether lipid fraction decreased quickly while that in the tetraether lipids increased simultaneously, without significant changes in the total radioactivity of ether lipids. These results strongly suggest that terbinafine inhibits the biosynthesis of tetraether lipids from a diether-type precursor lipid(s). The terbinafine treatment will be a tool for dissecting tetraether lipid biosynthesis in T. acidophilum.

Generation of a large, protonophore-sensitive proton motive force and pH difference in the acidophilic bacteria Thermoplasma acidophilum and Bacillus acidocaldarius.

The mechanism by which acidophilic bacteria generate and maintain their cytoplasmic pH close to neutrality was investigated. For this purpose we determined the components of proton motive force in the eubacterium Bacillus acidocaldarius and the archaebacterium Thermoplasma acidophilum. After correction for probe binding, the proton motive force of untreated cells was 190 to 240 mV between external pH 2 and 4. Anoxia diminished total proton motive force and the transmembrane pH difference by 60 to 80 mV. The protonophore 2,4-dinitrophenol abolished the total proton motive force almost completely and diminished the transmembrane pH difference by at least two units. However, even after correction for probe binding, protonophore-treated cells maintained a pH difference of approximately one unit.

ZETA-Potential and surface charge of Thermoplasma acidophila.

The surface charge density and the zeta-potential of Thermoplasma acidophila was estimated from microscopic electrophoresis experiments. The cells moved towards the positive electrode. The mobility remained constant from pH 2 to 5, and increased for pH values higher than 6. The mobility at pH 6 decreased dramatically with increased external Ca2+ concentration. At pH 2 and an ionic strength similar to that of the growth medium, the zeta-potential was about 8 mV, negative relative to the bulk medium; the surface charge density was 1360esu/cm-2 which corresponds to one elementary charge per 3500 A2.