Fish antifreeze proteins have unexpected applications

By Tammy Hardiman

In years to come, blood products may be stored for longer periods of time, a therapy for hypothermia may be developed, and a salmon with superior growth capabilities may become standard. These are just some of the breakthroughs that can be traced to the work of Dr. Garth Fletcher, professor of ocean sciences at Memorial's Ocean Sciences Centre (OSC).

For the past 24 years, Dr. Fletcher and his colleagues have been funded by the Natural Sciences and Engineering Research Council (NSERC) to study cold water adaptation in fish with an emphasis on the antifreeze proteins found in their blood. These vital proteins prevent ice crystals from growing in the blood when the water temperature is very low, thus preventing the fish from freezing.

Atlantic salmon do not possess antifreeze proteins and therefore are unable to be cultured in sub-zero waters off the Atlantic coast of Canada. In order to make salmon more freeze resistant, Dr. Fletcher injected antifreeze protein genes into salmon eggs. These genes were incorporated into the salmon's chromosomes and so far they have passed on through three successive generations of salmon.

However, as Dr. Fletcher explained, "The levels of antifreeze found in the salmon are low; thus, although the results are promising, more research is required before this salmon broodstock will be of value to industry."

This is not the only research ongoing to improve the performance of salmon for aquaculture. In 1990 Dr. Fletcher and his colleague, Dr. Choy Hew of the University of Toronto, developed a gene that contained a salmon growth hormone and antifreeze component. When Dr. Fletcher's team injected this gene into the salmon eggs, the resulting transgenic salmon grew four to six times faster than standard salmon.

Drs. Fletcher and Hew have recently been awarded a U.S. patent for this gene construct. "It is probably the first gene construct that has been patented in the production of a food animal," Dr. Fletcher noted.

Dr. Fletcher and Dr. Hew received another strategic NSERC grant to begin studies on the transfer of disease-resistant genes to salmon. With such ongoing research, the development of a superior salmon stock is within reach.

Antifreeze proteins isolated from Newfoundland fish sell for $5,000-$7,000 per gram, with one litre of fish blood yielding between one and four grams. Most of the blood serum comes from ocean pout fish at the Wesleyville hatchery, where people are trained to extract it. The serum is then sent to A/F Protein Canada where the proteins are isolated.

Dr. Fletcher is scientific adviser to A/F Protein Inc., a company based in Boston, Mass., which is marketing this technology internationally. Dr. Fletcher is also the president of a subsidiary company, A/F Protein Canada -- located in St. John's -- which has been created specifically to purify antifreeze proteins and assist in the marketing of transgenic fish. A/F Protein Canada is supported by funds from the Atlantic Canada Opportunities Agency and by the Industrial Research Assistance Program of the National Research Council.

A big market for the use of antifreeze proteins is in the preservation of blood platelets. Blood platelets are normally stored at room temperature for five days, after which they are no longer viable and are discarded. "[Protein antifreeze] could simplify or extend the shelf life of platelets for quite a long time and may mean less units collected," Dr. Fletcher explained. "It could change how the marketplace works for this product....of course it will have to go through regulatory agencies before it is used, but it is highly promising."

Further research has shown that the proteins protect mammalian eggs and other cells during cold or frozen storage, and the potential exists to use the proteins to protect human organs for transplantation. For example, rat livers infused with the proteins show a higher level of viability than non-infused ones. Dr. Fletcher says that much research is still needed before the new technology can be incorporated with the current practices, but the possibility is there for organs to be preserved for longer periods than is now possible.

Some companies in the frozen dessert industry are studying how antifreeze proteins modify the way that frozen foods recrystallize. Normally, when a product like ice cream melts and is frozen again it loses its smooth texture as ice crystals begin recrystallizing into bigger crystals. The antifreeze proteins can modify this process and prevent the crystals from growing. Such a breakthrough would be very lucrative since desserts, and ice cream in particular, are popular products.

There are still many hurdles to overcome before antifreeze proteins are fully understood and their applications become an everyday reality.