Research Tools : Fermentation


Creating ‘Designer’ Yeast Hybrids for Brewing and More

UW–Madison researchers have developed HyPr, a simple and efficient method for generating synthetic Saccharomyces hybrids without sporulation or modification of the nuclear genome.

Specifically, using the new method, induction of HO endonuclease expression by a promoter in two diploid cultures, followed by co-culture and subsequent double-drug selection, will produce hybrids at a rate approaching 1 out of 1,000 cells plated. Plasmids can then be easily cured or spontaneously lost to produce strains without genome modifications.

The resulting strains can be rapidly screened for plasmid loss, opening an efficient route towards meeting the Generally Recognized as Safe (GRAS) standards of the U.S. Department of Agriculture and FDA.

Yeast-Based Intein Platform for Drug Production

UW–Madison researchers have engineered non-self-cleaving Mxe GyrA inteins shown to significantly improve the production of fusion proteins from Saccharomyces cerevisiae. The novel inteins were developed through directed evolution, and they enhance fusion protein display (up to 3x) and secretion levels (up to 30x) compared to the wild type intein. The new yeast-based platform provides a robust alternative to bacterial intein expression systems.

Small Molecule Catalysts of Oxidative Protein Folding

UW–Madison researchers have synthesized novel catalysts for use in the production of high value recombinant proteins including antibodies and other biologics. The small molecules contain binding features that mimic PDI and are nearly optimal for catalyzing disulfide bond formation. Namely, pendant “R” groups provide affinity for the hydrophobic substrate (unfolded protein) but not the product (folded protein), enabling the catalysts to operate like a true enzyme.

The novel catalysts and method can be used in vivo or in vitro.

More Efficient Ethanol Production from Mixed Sugars Using Spathaspora Yeast

UW–Madison researchers have developed a method for producing ethanol using Spathaspora passalidarum yeast to ferment xylose or cellobiose, even when mixed with glucose.

The ethanol is converted from biomass or other lignocellulosic material from agricultural residues, fast-growing hardwoods and processing byproducts. Sugars, lignin and other components are first extracted from this feedstock using standard methods to form mixtures rich in different sugars. The mixture is contacted with a Spathaspora yeast cell under oxygen-limiting conditions suitable to allow the yeast to ferment a portion of the xylose and/or cellobiose into ethanol.

Thermostable Barley Alpha-Glucosidase for Improved Ethanol Production

UW-Madison researchers have developed a mutant barley alpha-glucosidase with increased thermal stability. They developed thermostable forms of the enzyme using site directed mutagenesis. Sites for mutagenesis were selected through comparisons with the sequences of other, more thermostable, alpha-glucosidase proteins.

Expression System and Fermentation Processes For Overexpression Of Holo-Acyl Carrier Protein

UW-Madison researchers have created eight bacterial expression plasmids from either E. coli or spinach, which express both apo-ACP and E. coli holo-ACP synthase, the enzyme responsible for the post-translational modification of ACP. Therefore, the plasmids allow completion of the required modification. The researchers' expression system enables the production of ACPs in high yields by using chemically defined minimal medium and specialized fermentation procedures.