Yeast strains
Inventors
Slininger, Patricia J. • Thompson, Stephanie R. • Balan, Venkatesh • Da Costa Sousa, Leonardo • Dale, Bruce • Shea-Andersh, Maureen A. • Dien, Bruce S.
Assignees
Board Of Turstees Of Michigan State University • Michigan State University MSU • US Department of Agriculture USDA
Publication Number
US-9297027-B1
Publication Date
2016-03-29
Expiration Date
2034-01-29
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Abstract
Several mutated strains of Scheffersomyces stipitis are generated by repetitive culturing of the parent strain on two types of concentrated hydrolyzates and with ethanol-challenged xylose-fed continuous culture. Isolates collected from various enriched populations are screened and ranked based on relative xylose uptake rate and ethanol yield. Ranking on hydrolyzates with and without nutritional supplementation is used to identify those isolates with best performance across diverse conditions.
Core Innovation
This invention relates to novel strains of Scheffersomyces stipitis that have enhanced capability of producing ethanol from diverse lignocellulosic hydrolyzates with diverse nutrient supplementation compared to the parent strain. These novel strains are generated through a method involving repetitive culturing on different media types such as low furfural concentration with high acetic acid and nitrogen (e.g., AFEX-CSH), media high in xylose with varying glucose content (e.g., PSGHL and SGH), and media containing substantial ethanol and high xylose concentration. Optionally, irradiation (such as UV) may be applied before or during culturing to enhance mutation and selection.
The problem addressed is that lignocellulosic biomass, while abundant, is challenging to deconstruct into fermentable sugars due to its fibrous structure and the presence of inhibitory byproducts like acetic acid and furan aldehydes created during pretreatment. Traditional industrial yeasts cannot ferment xylose nor survive in toxic concentrated hydrolyzates with high sugar concentrations needed for economical ethanol production. The naturally xylose-fermenting Scheffersomyces stipitis strain NRRL Y-7124, though superior to other yeasts, still requires improvement in sugar uptake rate, inhibitor tolerance, and ethanol production at high ethanol concentrations.
The invention provides a solution through selection and adaptation processes involving culturing the parent S. stipitis cells in one or multiple (up to four) different selective media that expose the strains to various stresses including inhibitors and ethanol. This selective pressure, combined with screening and ranking isolates for superior xylose uptake rates and ethanol yields across diverse media formulations, produces novel strains that demonstrate reduced diauxic lag during sugar transitions, faster growth, higher ethanol production (at least approximately 40 g/L), and tolerance to inhibitors in hydrolyzates. These strains are significantly improved over the parent strain in fermenting lignocellulosic hydrolyzates and are deposited under specified accession numbers.
Claims Coverage
The patent includes one independent claim relating to an isolated yeast strain and one independent claim relating to a method of producing ethanol using such strains. The main inventive features are identified as follows.
Isolated Scheffersomyces stipitis strain with enhanced ethanol production and reduced diauxic lag
An isolated S. stipitis strain that can produce at least 40 g/L ethanol when cultured on non-detoxified lignocellulosic hydrolyzate, and exhibits reduced diauxic lag during sugar transition from glucose to xylose compared to the parental S. stipitis strain NRRL Y-7124. The strain is selected from specific ARS patent deposits (NRRL Y-50871, Y-50859, Y-50862, Y-50864, Y-50861).
Reduced diauxic lag in the presence of ethanol
The isolated strain exhibits reduced diauxic lag during sugar transition from glucose to xylose in the presence of approximately 15 g/L or more ethanol.
Method of producing ethanol using the isolated S. stipitis strains
A method comprising growing at least one of the isolated S. stipitis strains on lignocellulosic hydrolyzate for a period effective to enable growth and fermentation of the hydrolyzate to ethanol.
The claims cover isolated yeast strains with improved ethanol production and reduced lag during sugar transitions under inhibitor and ethanol stress, as well as methods for ethanol production employing these strains.
Stated Advantages
The novel strains have enhanced capability to produce ethanol (at least approximately 40 g/L) from diverse lignocellulosic hydrolyzates compared to the parent strain.
They exhibit reduced diauxic lag during sugar transition from glucose to xylose, including in the presence of high ethanol concentrations (15-20 g/L or more).
They have faster initial growth, more rapid glucose and xylose uptake rates, and improved ethanol yield on xylose in the presence of inhibitors and under diverse nutritional environments.
The adapted strains are tolerant of various inhibitors present in hydrolyzates such as furan aldehydes, acetic acid, and ethanol, enabling fermentation at economically relevant ethanol concentrations.
Documented Applications
Production of ethanol by fermenting lignocellulosic biomass hydrolyzates comprising glucose and xylose sugars using the novel Scheffersomyces stipitis strains.
Use of the novel strains in media containing non-detoxified hydrolyzates with diverse nutrient supplementation, including AFEX-pretreated corn stover hydrolyzate, dilute acid-pretreated switchgrass hydrolyzate liquor (PSGHL), and enzyme-saccharified dilute acid-pretreated switchgrass hydrolyzate (SGH) with or without nutrient supplements.
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