Physicochemical, physiological, and engineering aspects of membrane derived vesicles from the extreme thermoacidophile Sulfolobus acidocaldarius.

摘要

Understanding the biochemical motifs that impart stability upon thermoacidophiles has implications in fundamental life sciences and in the engineering of stability in labile biocatalysts for the development industrial technologies.; The thermotropism of the model cell membrane was studied in an effort to address the gap between the simplified membrane models in the literature and what is actually going on in the cells. A novel, accurate cell model of a thermoacidophilic cell membrane was developed. The sub gel phase thermotropism was studied using differential scanning calorimetry (DSC). This work demonstrated that DSC is not applicable to observing phase transitions in the improved model because of the large amount of protein present in the membrane.; The viability of cell cultures incubated at low temperatures in the presence of transmembrane pH gradients was followed to determine if there were subgel phase transitions in the membrane that impacted the viability of the thermoacidophile at low temperature. Based on this work there is no deviation from expected cell death rate. It was concluded that no phase transition is present within two weeks under these conditions.; Amyloglucosidase, a labile, soluble enzyme with diverse applications in industry, was entrapped in mesophilic-lipid liposomes, and an extension of activity under optimal conditions of temperature and pH was observed. The upper limits of the active window of pH and temperature for this enzyme were explored. This enzyme was found to be unsuitable for studies at 75°C. Entrapment and evaluation of amyloglucosidase in the thermoacidophilic model membrane demonstrated that the vesicle is an effective vehicle for the entrapment and recovery of the biocatalyst. The reaction kinetics of the enzyme were characterized in the free and entrapped state---both in the thermoacidophilic and the mesophilic systems. Kinetic parameters and substrate mass transfer were determined in each system.