ENHANCED SURVIVAL OF YEAST EXPRESSING AN ANTIFREEZE GENE ANALOG AFTER FREEZING

Yeast, like most organisms, survives poorly under freezing conditions. It has been proposed that after rapid cooling yeast suffers a loss in viability from the recrystallization of intracellular ice. Antifreeze proteins found in the blood of certain polar fishes have been shown to be potent inhibitors of ice recrystallization at very low concentrations. We have examined the feasibility of protecting rapidly cooled yeast cells from freezing damage by inhibiting the recrystallization of intracellular ice through in vivo expression of an antifreeze analogue gene. A chemically synthesized gene encoding a protein similar to but differing from the antifreeze proteins of the fish Pseudopleuronectes americanus (winter flounder) was genetically fused to the 3′ end of a truncated staphylococcal Protein A gene. When the fused gene was expressed in the budding yeast Saccharomyces cerevisiae, its cells were shown to produce a new chimeric protein that inhibited the recrystallization of ice in vitro. Yeast cells expressing the chimeric antifreeze protein showed a twofold increase in survival after rapid freezing (95 °C/min to -196 °C) and moderate rates of warming (26 to 64 °C/min) compared to cells lacking the chimeric protein. © 1991.