Flowers of the hop plant provide both bitterness and “hoppy” flavor to beer. Hops are, however, both a water and energy intensive crop and vary considerably in essential oil content, making it challenging to achieve a consistent hoppy taste in beer. Here, we report that brewer’s yeast can be engineered to biosynthesize aromatic monoterpene molecules that impart hoppy flavor to beer by incorporating recombinant DNA derived from yeast, mint, and basil. Whereas metabolic engineering of biosynthetic pathways is commonly enlisted to maximize product titers, tuning expression of pathway enzymes to affect target production levels of multiple commercially important metabolites without major collateral metabolic changes represents a unique challenge. By applying state-of-the-art engineering techniques and a framework to guide iterative improvement, strains are generated with target performance characteristics. Beers produced using these strains are perceived as hoppier than traditionally hopped beers by a sensory panel in a double-blind tasting.
Whereas domestication of livestock, pets, and crops is well documented, it is still unclear to what extent microbes associated with the production of food have also undergone human selection and where the plethora of industrial strains originates from. Here, we present the genomes and phenomes of 157 industrial Saccharomyces cerevisiae yeasts. Our analyses reveal that today’s industrial yeasts can be divided into five sublineages that are genetically and phenotypically separated from wild strains and originate from only a few ancestors through complex patterns of domestication and local divergence. Large-scale phenotyping and genome analysis further show strong industry-specific selection for stress tolerance, sugar utilization, and flavor production, while the sexual cycle and other phenotypes related to survival in nature show decay, particularly in beer yeasts. Together, these results shed light on the origins, evolutionary history, and phenotypic diversity of industrial yeasts and provide a resource for further selection of superior strains. PAPERCLIP.
During the beer brewing process, bitter tasting cis and trans iso-α-acids are generated from the precursor α-acids found in hops. The absolute configurations of the α-acid (-)-humulone and several of its derivatives have now been elucidated by X-ray crystallography, thus resolving decades of confusion over the humulone isomerization mechanism.
Intracellular triacylglycerol (TAG) is a ubiquitous energy storage lipid also involved in lipid homeostasis and signaling. Comparatively, little is known about TAG’s role in other cellular functions. Here we show a pro-longevity function of TAG in the budding yeast Saccharomyces cerevisiae. In yeast strains derived from natural and laboratory environments a correlation between high levels of TAG and longer chronological lifespan was observed. Increased TAG abundance through the deletion of TAG lipases prolonged chronological lifespan of laboratory strains, while diminishing TAG biosynthesis shortened lifespan without apparently affecting vegetative growth. TAG-mediated lifespan extension was independent of several other known stress response factors involved in chronological aging. Because both lifespan regulation and TAG metabolism are conserved, this cellular pro-longevity function of TAG may extend to other organisms.
Beyond its classical biotechnological applications such as food and beverage production or as a cell factory, the yeast Saccharomyces cerevisiae is a valuable model organism to study fundamental mechanisms of cell response to stressful environmental changes. Acetic acid is a physiological product of yeast fermentation and it is a well-known food preservative due to its antimicrobial action. Acetic acid has recently been shown to cause yeast cell death and aging. Here we shall focus on the molecular mechanisms of S. cerevisiae stress adaptation and programmed cell death in response to acetic acid. We shall elaborate on the intracellular signaling pathways involved in the cross-talk of pro-survival and pro-death pathways underlying the importance of understanding fundamental aspects of yeast cell homeostasis to improve the performance of a given yeast strain in biotechnological applications.
Many crops display differential geographic phenotypes and sensorial signatures, encapsulated by the concept of terroir. The drivers behind these differences remain elusive, and the potential contribution of microbes has been ignored until recently. Significant genetic differentiation between microbial communities and populations from different geographic locations has been demonstrated, but crucially it has not been shown whether this correlates with differential agricultural phenotypes or not. Using wine as a model system, we utilize the regionally genetically differentiated population of Saccharomyces cerevisiae in New Zealand and objectively demonstrate that these populations differentially affect wine phenotype, which is driven by a complex mix of chemicals. These findings reveal the importance of microbial populations for the regional identity of wine, and potentially extend to other important agricultural commodities. Moreover, this suggests that long-term implementation of methods maintaining differential biodiversity may have tangible economic imperatives as well as being desirable in terms of employing agricultural practices that increase responsible environmental stewardship.
Benzylisoquinoline alkaloids (BIAs) are a diverse family of plant-specialized metabolites that include the pharmaceuticals codeine and morphine and their derivatives. Microbial synthesis of BIAs holds promise as an alternative to traditional crop-based manufacturing. Here we demonstrate the production of the key BIA intermediate (S)-reticuline from glucose in Saccharomyces cerevisiae. To aid in this effort, we developed an enzyme-coupled biosensor for the upstream intermediate L-3,4-dihydroxyphenylalanine (L-DOPA). Using this sensor, we identified an active tyrosine hydroxylase and improved its L-DOPA yields by 2.8-fold via PCR mutagenesis. Coexpression of DOPA decarboxylase enabled what is to our knowledge the first demonstration of dopamine production from glucose in yeast, with a 7.4-fold improvement in titer obtained for our best mutant enzyme. We extended this pathway to fully reconstitute the seven-enzyme pathway from L-tyrosine to (S)-reticuline. Future work to improve titers and connect these steps with downstream pathway branches, already demonstrated in S. cerevisiae, will enable low-cost production of many high-value BIAs.
- Proceedings of the National Academy of Sciences of the United States of America
- Published about 8 years ago
Saccharomyces cerevisiae is one of the most important model organisms and has been a valuable asset to human civilization. However, despite its extensive use in the last 9,000 y, the existence of a seasonal cycle outside human-made environments has not yet been described. We demonstrate the role of social wasps as vector and natural reservoir of S. cerevisiae during all seasons. We provide experimental evidence that queens of social wasps overwintering as adults (Vespa crabro and Polistes spp.) can harbor yeast cells from autumn to spring and transmit them to their progeny. This result is mirrored by field surveys of the genetic variability of natural strains of yeast. Microsatellites and sequences of a selected set of loci able to recapitulate the yeast strain’s evolutionary history were used to compare 17 environmental wasp isolates with a collection of strains from grapes from the same region and more than 230 strains representing worldwide yeast variation. The wasp isolates fall into subclusters representing the overall ecological and industrial yeast diversity of their geographic origin. Our findings indicate that wasps are a key environmental niche for the evolution of natural S. cerevisiae populations, the dispersion of yeast cells in the environment, and the maintenance of their diversity. The close relatedness of several wasp isolates with grape and wine isolates reflects the crucial role of human activities on yeast population structure, through clonal expansion and selection of specific strains during the biotransformation of fermented foods, followed by dispersal mediated by insects and other animals.
It has been speculated that the partial pressure of carbon dioxide (pCO2) in shelf waters may lag the rise in atmospheric CO2. Here, we show that this is the case across many shelf regions, implying a tendency for enhanced shelf uptake of atmospheric CO2. This result is based on analysis of long-term trends in the air-sea pCO2 gradient (ΔpCO2) using a global surface ocean pCO2 database spanning a period of up to 35 years. Using wintertime data only, we find that ΔpCO2 increased in 653 of the 825 0.5° cells for which a trend could be calculated, with 325 of these cells showing a significant increase in excess of +0.5 μatm yr-1 (p < 0.05). Although noisier, the deseasonalized annual data suggest similar results. If this were a global trend, it would support the idea that shelves might have switched from a source to a sink of CO2 during the last century.
Morphinan alkaloids are the most powerful narcotic analgesics currently used to treat moderate to severe and chronic pain. The feasibility of morphinan synthesis in recombinant Saccharomyces cerevisiae starting from the precursor (R,S)-norlaudanosoline was investigated. Chiral analysis of the reticuline produced by the expression of opium poppy methyltransferases showed strict enantioselectivity for (S)-reticuline starting from (R,S)-norlaudanosoline. In addition, the P. somniferum enzymes salutaridine synthase (PsSAS), salutaridine reductase (PsSAR) and salutaridinol acetyltransferase (PsSAT) were functionally co-expressed in S. cerevisiae and optimization of the pH conditions allowed for productive spontaneous rearrangement of salutaridinol-7-O-acetate and synthesis of thebaine from ®-reticuline. Finally, we reconstituted a 7-gene pathway for the production of codeine and morphine from ®-reticuline. Yeast cell feeding assays using ®-reticuline, salutaridine or codeine as substrates showed that all enzymes were functionally co-expressed in yeast and that activity of salutaridine reductase and codeine-O-demethylase likely limit flux to morphine synthesis. The results of this study describe a significant advance for the synthesis of morphinans in S. cerevisiae and pave the way for their complete synthesis in recombinant microbes.