Protein keeps cold fish from becoming frozen flounder, new study shows

'Hyperactive' antifreeze protein has eluded researchers for more than 30 years.

(Kingston, ON) – A surprising discovery by Queen's researchers helps explain why fish swimming in icy sea water don't freeze.

The team, led by Biochemistry Professor Peter Davies, has identified a new "antifreeze" protein found in the blood of winter flounder enabling the fish to withstand temperatures as low as -1.9 degrees Celsius: the freezing point of sea water. The antifreeze plasma proteins (AFPs) do this by binding irreversibly to ice crystals and preventing them from growing.

Until now, it has been a mystery how these fish survive in polar oceans, since the previously identified "type I" AFP associated with winter flounder only provides 0.7oC of freezing point depression, which in combination with blood solutes, only protects the fish down to -1.5 degrees Celsius.

"This finally explains the 'critical gap' of 0.4 degrees," says Dr. Davies, a Queen's Canada Research Chair in Protein Engineering. "The winter flounder has been studied extensively by a number of laboratories over the past 30 years, but this antifreeze protein escaped everyone's notice. We're excited to have found it."

The research, conducted with Christopher Marshall from Queen's Department of Biochemistry and Garth Fletcher from the Ocean Sciences Centre at Memorial University, is published today in the journal Nature.

The team used a process called ice affinity purification to identify the new protein. "When you grow a 'popsicle' of ice in the presence of these proteins, the AFPs bind to the ice and become included, while other proteins are excluded," explains Mr. Marshall. "Lemon-shaped ice crystals that differed significantly from the hexagon-shaped crystals obtained with type I AFPs told us that we were dealing with an unknown antifreeze protein."

The new protein is extraordinarily active in comparison with other fish antifreeze proteins. At room temperature and at low pH values, however, it loses all activity – perhaps explaining why it remained undetected for three decades. "Prior to this we had only found such hyperactive antifreeze proteins in insects," says Dr. Davies.

Being able to control the growth of ice crystals could have a number of bio-technological and medical applications, the researchers suggest.

AFPs have been tested in the storage of organs and blood products for transplantation, where they offer protection against freezing, improving viability and extending maximum storage periods. They have also been applied in cryosurgery, a technique in which tumor cells are killed by freezing, because AFPs modify the shape of ice crystals into more destructive spicules.

This finding also opens the possibility of transferring genes from winter flounder into salmon, for example, to make them more freeze-resistant for fish farming, or into crops to make them more frost-resistant to extend their growing season. These applications could be realized with concentrations of hyperactive AFPs 10 to 100-fold lower than would be required with the previously discovered fish AFPs.

12-may-2004