Abstract

Strains of Vitreoscilla stercoraria were isolated from the environment and characterised. Cell width, motility and requirement of each strain for sodium chloride were investigated. Two strains were selected for further study and the effect of monensin and FCCP on growth of the strains was investigated. One strain of Vitreoscilla (LB13) was chosen for further study, cells from strain LB 13 were found to be 1.38 µm ± 0.041 (± 1 SEM, n=10) wide, were motile by gliding and had an optimum requirement for sodium chloride for growth of 43 mM. The organism was grown in batch culture and respiratory membranes were isolated. Cytochrome bo was extracted from the respiratory membranes and further purification was achieved using column chromatography. A yield of 6.71 % was achieved with a purification factor of 18.5. The light sensitivity of Vitreoscilla stercoraria was investigated. Two strains of Vitreoscilla (LB13 and C1) were shown to be highly sensitive to UV-A (320-400 nm) with an LD50 of less than 20 kJmˉ². Superoxide dismutase and catalase were shown to provide protection from the effect of UV-A during exposure, either separately of together, indicating an involvement of reactive oxygen species. A photo-insensitive strain (LB13A) was isolated during an exposure experiment and originated from a culture of LB13. The possible sodium pumping activity of cytochrome bo from two strains of Vitreoscilla (LB13 and C1) was investigated. The Vmax of decylubiquinol oxidation by respiratory membranes from LB13 and C 1 were calculated and found to be 0.96 nmol sˉ¹ mg-1 and 13.33 nmol sˉ¹ mgˉ¹ respectively. The Km of decylubiquinol oxidation by LB13 membranes was found to be 9.8 µM. Quinol oxidation activity was tested for dependence on sodium chloride in both respiratory membranes and in the purified enzyme. No stimulation of activity was shown with either strain using decylubiquinol, duroquinol or menadiol as substrates. Given the lack of sodium sensitivity it is unlikely that the enzyme pumps sodium.

Document Type

Thesis

Publication Date

2003

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