Yeast Biology
Brewer’s yeast are living organisms. Just two species of yeast — Saccharomyces cerevisiae (ale yeast) and S. pastorianus (lager yeast) — are used in the overwhelming majority of beer fermentations.
In the wild, yeast live on rotting fruit, tree sap or any other source of simple sugars. Most yeast species consume simple sugars as their sole carbon source, although some can metabolize organic acids or alcohol. Yeast can break down sugars via the Krebs Cycle, when oxygen is present. In the absence of oxygen, most species can ferment sugar, producing alcohol. The preference of most yeast to use aerobic respiration, when oxygen is present, is called the Pasteur Effect.
Brewer’s yeast is unusual in that, in the presence of both sugar and oxygen, it ferments sugars. This sometimes called the Crabtree Effect. It does this despite the fact that aerobic respiration yields far more energy than alcoholic fermentation. In wild strains of S. cerevisiae, both oxygen and alcohol are present soon after they colonize a piece of fruit. When the alcohol level reaches a certain point, the yeast do something remarkable . . . they begin to consume the alcohol they produced. Alcohol taken in by the yeast is converted to acetaldehyde, then acetate, then acetyl phosphate and finally acetyl CoA, before being fed into the Krebs Cycle. In brewery fermentations, this does not happen to any appreciable degree because oxygen is excluded from the beer (and hence the Krebs Cycle cannot function). A group in Japan did find, however, that yeast with a knocked out ADH2 gene — which codes for the enzyme that converts alcohol to acetaldehyde — produced beer with lower levels of acetaldehyde.
Why would S. cerevisiae ferment sugars, only to consume a part of the ethanol they produce later? The answer is that alcohol is toxic to most other microorganisms. Brewer’s yeast initially begins producing alcohol to poison the environment for other species. Once it has crowded out most of it’s rivals, it takes in some of the alcohol it has made and degrades it via the Krebs Cycle.
Of the 1,500 species of yeast — and the tens of millions of bacterial species — only two are good brewing species, S. cerevisiae (ale yeast) and S. pastorianus (lager yeast). (If you include wine in the conversation, you can add S. bayanus to the list.) But what makes a yeast suitable for use in brewing? How did S. cerevisiae and S. pastorianus acquire the characteristics that make them good at brewing? Scientists in the last ten years have found out some of the details. S. cerevisiae is not just a favorite of brewers, its also a favorite of biologists. In fact, S. cerevisiae has been a model organism for many years. As such, it was the first eukaryote to be completely sequenced (back in 1996). Just one year later, however, scientists made an astounding discovery.
Whole Genome Duplication
What scientists found was that S. cerevisiae had undergone a whole genome duplication 100 million years ago (MYA). The primary evidence for this comes from looking at the modern S. cerevisiae genome and comparing it to the genome of related species, such as Kluyveromyces lactis. Kluyveromyces is a yeast with 8 chromosomes and its chromosomes map onto the S. cerevisiae genome in a 1:2 fashion. For each Kluyveromyces chromosome, there are two Saccharomyces chromosomes that have the same genes, in the same order. But, there’s a twist. Many of the genes on the two Saccharomyces chromosomes are missing, but in a complementary manner. One way to visualize this is to think of a hypothetical chromosome with 10 genes, named (for convenience) 1 through 10, in that order. The Kluyveromyces chromosome would have those 10 genes, in that order. The two Saccharomyces chromosomes might contain the genes 1, 2, 3, 7, 8, 9 and 3, 4, 5, 6, 10, respectively (and in those orders). The two Saccharomyces chromosomes contain all of the genes found in the Kluyveromyces chromosome, just spread out over two chromosomes — with one interesting twist.
You may have noticed that there’s a “3” in both of our example chromosomes. This illustrates something else researchers found. Although most of the duplicate genes had been deleted (or inactivated) in the modern Saccharomyces genome, about 10 percent of the genes were present in two copies. Of the two copies, one was always similar to the corresponding gene in Kluyveromyces. The other frequently showed that its sequence had diverged from other copy. And for brewers, the interesting aspect of this is that most of the doubly-retained genes deal with sugar metabolism. And the kicker is, the timing of the genome duplication (100 MYA) corresponds with the rise of the Angiosperms — plants that produce flowers and fleshy, sugary fruits.
Duplication of Alcohol Dehydrogenase
About 80 MYA, the ancestor to modern S. cerevisiae experienced a small duplication event, involving only part of a single chromosome. This created two copies of the enzyme alcohol dehydrogenase, ADH1 and ADH2. ADH1 catalyses the formation of alcohol from acetaldehyde, whereas ADH2 catalyses the reverse reaction. The presence of two different ADH enzymes, with opposing functions is, as you have probably guessed, part of the basis of how S. cerevisiae can consume alcohol, as described above.
The sequences of ADH1 and ADH2 are very similar, and ADH1 in S. cerevisiae is similar to one of the ADH enzymes in Kluyveromyces (which has independently duplicated its ADH genes twice, yielding four genes). From this, scientists inferred that ADH2 was the gene with “new” function and that the ancestral ADH gene functioned to produce, not consume, alcohol. They confirmed this by comparing ADH1 (from S. cerevisiae) to the appropriate ADH gene in Kluyveromyces and inferring all the most likely ancestral sequences. They then synthesized these enzymes and tested their kinetics (how they perform, basically.) Most of the “resurrected” enzymes had some function and in most it was predominantly to produce alcohol from acetaldehyde.
The Origin of Lager Yeast
Lager yeast, S. pastorianus, ferments at significantly colder temperatures than ale yeast. It has long been known that S. pastorianus has a different set of chromosomes than S. cerevisiae, and two fundamentally different types of lager yeast exist. Recently, researchers found out the source of this difference. As it turns out, one type of lager yeast is a hybrid between S. cerevisiae and S. bayanus, a yeast that can ferment at much lower temperatures than S. cerevisiae. The other type of lager yeast is a second hybrid between S. cerevisiae and S. bayanus. In both cases, the S. bayanus genome is almost entirely retained, while S. cerevisiae-derived sequences have mostly been lost. By comparing the remaining cerevisiae-derived sequences in S. pastorianus to wild species of S. cerevisiae and also strains that ferment wine, sake and ales, researchers concluded that, in both cases, the cerevisiae-derived sequences in lager yeast came from an ale strain. However, in each case, the ale strain was different. This is not surprising since it has always been expected that S. pastorianus arose in a brewery.
Tastes Great, Less Phenols
One final step in brewer’s yeast evolution has been very important, but is incredibly simple to explain. Most wild species of yeast produce off flavors and aromas, with 4-vinyl-guiacol (a phenolic compound) being the most prevalent. In brewer’s yeast, this gene (named POF) has apparently been deleted.
So next time you brew, think of all the duplications, deletions and divergences that your yeast’s ancestors had to go through to bring you your tasty fermented grain beverage — and raise a glass to the scientists who are working on finding out the whole story.
