Diter von Wettstein, retired at age 67 from the Carlsberg Research Center in Copenhagen, could have rested on his many laurels; instead, he came to WSU to become the R.A. Nilan distinguished professor of barley research and education.

"I plan to use the techniques for genetic transformation of barley to improve barley in the Northwest," says von Wettstein. These techniques, however, take time.

Grain is the barley's fruit, and each grain contains but one seed, the embryo that will grow into a new barley plant. Most of the rest of the grain—about 80 percent of it—is endosperm and contains nutrients for the embryo in the form of starches and proteins.

When the barley embryo germinates or sprouts, it sends a hormone message to the cells surrounding the endosperm. These cells then produce enzymes that move into the endosperm and break down its starches and proteins into sugars and amino acids—smaller molecules the embryo and developing seedling can use.

Brewers take advantage of this, harvesting the sugars as food for the yeast that will make beer out of the barley and hops. They allow grains to undergo controlled germination during the process known as "malting."

Currently the brewer orchestrates a balancing act between two of the enzymes that are produced and delivered into the endosperm. One is beta-glucanase. It breaks down the walls that divide the endosperm into starch-filled compartments. The other breaks down starches into small sugars.

Ideally, malting would leave all the walls broken down, for those that remain become viscous during the next stage in brewing, a problem that prevents the use of most barley varieties for malting. Yet if the grain germinates long enough for all the walls to be broken down, says von Wettstein, some of the starches will be broken down into sugars, and the developing seedling will use them as it grows. Those sugars won't be available as food for the yeast.

Hence the balancing act. The brewer must stop the malting when enough of the walls have been broken down, but not too many. He stops it with heat, which destroys the beta-glucanase.

Hoping to overcome this unwanted effect of heat, Von Wettstein decided to genetically engineer barley by introducing into it a gene that will enable the barley to make a beta-glucanase that will survive the heat. Then the enzyme can continue to break down the walls of the endo-sperm after germination has been stopped, clearing up the viscosity problem.

Obviously, this would help brewers and those who grow barley for them. But the same technology—of genetically engineering barley to make a foreign molecule when and where you want it—is also being used to develop medicines. Von Wettstein is collaborating on a project that will enable barley to produce a protein that will prevent degradation of the lining of the lungs. Seventy-thousand to 100,000 people in the United States, and many more worldwide, need this protein to stay alive, for their bodies don't make it. Yet it's available only in limited amounts and is expensive, for the only current source is human blood.

While it's true that the same protein might be made in yeast and bacteria, each of which is used to produce other medicinal molecules, each has problems. Making large amounts of the protein in yeast would be expensive. It would take a lot of sugar, for one thing.

Producing proteins in bacteria doesn't always produce proteins that work in humans. Humans, other animals, and plants add sugars to their newly made proteins. Bacteria don't. And these sugars are necessary for the long-term activity of the lung protein.

In fact, the addition of sugars poses an interesting long-term question for this project. Although both plants and humans add sugars, they do so differently. People add an extra piece to theirs. Von Wettstein doesn't know if what the plant does will be sufficient, or whether that extra piece will have to be added after the protein is isolated from the endosperm and purified.

Like the heat-stable beta-glucanase, the lung protein is made in response to a signal from the embryo at the time of germination and results in the protein being made and secreted into the endosperm. "Then industrial malting procedures can be used to extract this protein from the barley," says von Wettstein.

But barley grains can also be genetically engineered to produce and store other compounds in the endosperm and do so during the growth of the grain, not just at germination.

Barley is used to make animal feed, but is not germinated in the process. If the walls in the endosperm aren't broken down, they cause a problem for chickens, in particular, and pigs. So you need a beta-glucanase that works before germination. In addition, the process of making feed pellets produces heat, and chicken feed must be heated to destroy salmonella. This means you also need a heat-stable beta-glucanase, the same enzyme, but with different directions on when it's produced.

Creating the heat-stable beta-glucanase was a combination of good science, intelligence, and good luck. At the time von Wettstein began looking into the project, he had help from a colleague who was knowledgeable about beta-glucanases. His colleague knew that the same enzyme from two different bacterial species was somewhat more stable than that of barley and would work in the barley.

Their question: "Where does the heat stability reside?" A convenient cleavage site in the gene for each of the two enzymes lets them combine the front end of one bacteria's enzyme with the back end of the other. The new enzyme made from this hybrid gene was much more heat-stable than that from either bacteria. "This is the first case of hybrid vigor by recombination between two forms of the same gene," says von Wettstein, who also admits that this step—the convenient cleavage site—was the pure-luck part of the process.

Of course they tried many more combinations of parts from both enzymes, and eventually found one that could withstand four hours of heat. The original enzyme from barley could last four minutes. They also found the answer to their question of where the heat stability was—in the structural relationship between the front and back ends of the protein. Then they made other genetic changes and produced an enzyme that was heat-stable and worked well. Since then, it's been introduced into barley. The first Palouse field was planted with the new seeds in the summer of 1996, and by the end of the summer of 1997, von Wettstein should have enough genetically engineered barley to do malting experiments.

—Mary Aegerter