Many organisms that survive extreme dehydration (including, for example, yeast cells, seeds, fungal spores, and certain microscopic animals) contain large quantities of sugars, particularly the disaccharides sucrose or trehalose. Previous studies have shown that subcellular fractions such as labile enzymes and membranes can be preserved intact if they are dried in the presence of these sugars. If they are dried without the sugars these molecular assemblages are completely and irreversibly degraded. This phenomenon has been used to develop means for preserving proteins, membranes, and liposomes in the absence of water, leading to several patents that are in use in industry. The current project extends the information about subcellular fractions to intact cells. Fourier transform infrared spectroscopy is used to study the state of membranes in intact yeast cells that have been dried in ways that result in accumulation of various amounts of trehalose. These studies are designed to test the hypothesis that trehalose stabilizes membranes in intact cells in much the same ways as it does in isolated membranes. A trehalose-dependent phase diagram will be produced for the cells that should not only test this hypothesis in detail, but will also be of immediate practical use in the yeast industry in effecting increased survival of dehydration. A means will be sought to introduce trehalose into cells in the dry state that would not normally survive dehydration. The route chosen is to use a transporter for trehalose that is found in yeasts. Molecular biology techniques will be used to clone the gene for this transporter. Mutants have already been developed by collaborators in this research that lack this gene. The aim is to obtain the gene by complementation of these mutants. If successful, the gene will be introduced into other cells in subsequent research.
