Outer and Inner Membrane Preparations of Extreme Thermophile and Their Physico-Chemical Properties

Abstract

Outer and inner (cytoplasmic) membranes were partially purified from the gram negative extremely thermophilic bacteria, Thermus thermophilus HB-8 by sucrose density gradient centrifugation. In spite of our efforts to separate them, the inner membrane fraction contained some outer membrane components as determined by enzyme assay and electrophoresis.
When studied by 5DS spin labeling, the outer membranes showed a larger 2T₁₁ value (lower fluidity) than the inner membranes, although the fatty acid compositions were similar. The inner membranes of the cells cultured at higher temperature showed a larger 2T₁₁ value than the cells cultured at lower temperature. A similar phenomenon was observed with the TEMPO parameter of liposomal membranes. The upper break point (T_h) of the inner membranes observed by spin labeling was slightly lower than the culture temperature of the cells, and the lower break point (T₁) corresponded well to the lowest temperature limit of growth.
The calorimetric heating curve of the inner membranes had a broader temperature range of transition than that of the liposomal membranes. The transition temperature observed by calorimetry seems to reflect the melting properties of the membrane lipids, while fatty acid spin probe probably reports the local environment of the membrane, which is more directly related to its bilogical function.