scholarly journals 3′ Truncation of theGPD1Promoter in Saccharomyces cerevisiae for Improved Ethanol Yield and Productivity

2013 ◽  
Vol 79 (10) ◽  
pp. 3273-3281 ◽  
Author(s):  
Wen-Tao Ding ◽  
Guo-Chang Zhang ◽  
Jing-Jing Liu
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Yield ◽  
Research Effort ◽  
Final Concentration ◽  
Ethanol Productivity ◽  
Promoter Strength ◽  
Content Type ◽  
Very High Gravity ◽  
New Strategy ◽  
Glycerol Formation

ABSTRACTGlycerol is a major by-product in bioethanol fermentation by the yeastSaccharomyces cerevisiae, and decreasing glycerol formation for increased ethanol yield has been a major research effort in the bioethanol field. A new strategy has been used in the present study for reduced glycerol formation and improved ethanol fermentation performance by finely modulating the expression ofGPD1in the KAM15 strain (fps1ΔpPGK1-GLT1 gpd2Δ). TheGPD1promoter was serially truncated from the 3′ end by 20 bp to result in a different expression strength ofGPD1. The two engineered promoters carrying 60- and 80-bp truncations exhibited reduced promoter strength but unaffected osmostress response. These two promoters were integrated into the KAM15 strain, generating strains LE34U and LE35U, respectively. The transcription levels of LE34U and LE35U were 37.77 to 45.12% and 21.34 to 24.15% of that of KAM15U, respectively, depending on osmotic stress imposed by various glucose concentrations. In very high gravity (VHG) fermentation, the levels of glycerol for LE34U and LE35U were reduced by 15.81% and 30.66%, respectively, compared to KAM15U. The yield and final concentration of ethanol for LE35U were 3.46% and 0.33% higher, respectively, than those of KAM15U. However, fermentation rate and ethanol productivity for LE35U were reduced. On the other hand, the ethanol yield and final concentration for LE34U were enhanced by 2.28% and 2.32%, respectively, compared to those of KAM15U. In addition, a 2.31% increase in ethanol productivity was observed for LE34U compared to KAM15U. These results verified the feasibility of our strategy for yeast strain development.

scholarly journals The metabolic costs of improving ethanol yield by reducing glycerol formation capacity under anaerobic conditions in Saccharomyces cerevisiae

2013 ◽  
Vol 12 (1) ◽  
pp. 29 ◽  
Author(s):  
Julien Pagliardini ◽  
Georg Hubmann ◽  
Sandrine Alfenore ◽  
Elke Nevoigt ◽  
Carine Bideaux ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Yield ◽  
Anaerobic Conditions ◽  
Metabolic Costs ◽  
Glycerol Formation

scholarly journals Cofermentation of Cellobiose and Galactose by an Engineered Saccharomyces cerevisiae Strain

2011 ◽  
Vol 77 (16) ◽  
pp. 5822-5825 ◽  
Author(s):  
Suk-Jin Ha ◽  
Qiaosi Wei ◽  
Soo Rin Kim ◽  
Jonathan M. Galazka ◽  
Jamie Cate ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Neurospora Crassa ◽  
Plant Biomass ◽  
Ethanol Yield ◽  
Content Type ◽  
Marine Plant ◽  
Cellodextrin Transporter ◽  
Engineered Saccharomyces Cerevisiae

ABSTRACTWe demonstrate improved ethanol yield and productivity through cofermentation of cellobiose and galactose by an engineeredSaccharomyces cerevisiaestrain expressing genes coding for cellodextrin transporter (cdt-1) and intracellular β-glucosidase (gh1-1) fromNeurospora crassa. Simultaneous fermentation of cellobiose and galactose can be applied to producing biofuels from hydrolysates of marine plant biomass.

scholarly journals Directed Evolution of Xylose Isomerase for Improved Xylose Catabolism and Fermentation in the Yeast Saccharomyces cerevisiae

2012 ◽  
Vol 78 (16) ◽  
pp. 5708-5716 ◽  
Author(s):  
Sun-Mi Lee ◽  
Taylor Jellison ◽  
Hal S. Alper
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Production ◽  
Directed Evolution ◽  
Ethanol Yield ◽  
Xylose Isomerase ◽  
Mutant Enzyme ◽  
Xylose Utilization ◽  
Xylose Consumption ◽  
Content Type ◽  
Consumption Rates

ABSTRACTThe heterologous expression of a highly functional xylose isomerase pathway inSaccharomyces cerevisiaewould have significant advantages for ethanol yield, since the pathway bypasses cofactor requirements found in the traditionally used oxidoreductase pathways. However, nearly all reported xylose isomerase-based pathways inS. cerevisiaesuffer from poor ethanol productivity, low xylose consumption rates, and poor cell growth compared with an oxidoreductase pathway and, additionally, often require adaptive strain evolution. Here, we report on the directed evolution of thePiromycessp. xylose isomerase (encoded byxylA) for use in yeast. After three rounds of mutagenesis and growth-based screening, we isolated a variant containing six mutations (E15D, E114G, E129D, T142S, A177T, and V433I) that exhibited a 77% increase in enzymatic activity. When expressed in a minimally engineered yeast host containing agre3knockout andtal1andXKS1overexpression, the strain expressing this mutant enzyme improved its aerobic growth rate by 61-fold and both ethanol production and xylose consumption rates by nearly 8-fold. Moreover, the mutant enzyme enabled ethanol production by these yeasts under oxygen-limited fermentation conditions, unlike the wild-type enzyme. Under microaerobic conditions, the ethanol production rates of the strain expressing the mutant xylose isomerase were considerably higher than previously reported values for yeast harboring a xylose isomerase pathway and were also comparable to those of the strains harboring an oxidoreductase pathway. Consequently, this study shows the potential to evolve a xylose isomerase pathway for more efficient xylose utilization.

scholarly journals Improving Acetic Acid and Furfural Resistance of Xylose-Fermenting Saccharomyces cerevisiae Strains by Regulating Novel Transcription Factors Revealed via Comparative Transcriptomic Analysis

2021 ◽  
Vol 87 (10) ◽  
Author(s):  
Bo Li ◽  
Li Wang ◽  
Ya-Jing Wu ◽  
Zi-Yuan Xia ◽  
Bai-Xue Yang ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Acetic Acid ◽  
Transcription Factors ◽  
Ethanol Yield ◽  
Regulation Mechanism ◽  
Lignocellulosic Hydrolysate ◽  
Inhibitor Tolerance ◽  
Xylose Consumption ◽  
Content Type ◽  
Beneficial Effects

ABSTRACT Acetic acid and furfural are the two prevalent inhibitors coexisting with glucose and xylose in lignocellulosic hydrolysate. The transcriptional regulations of Saccharomyces cerevisiae in response to acetic acid (Aa), furfural (Fur), and the mixture of acetic acid and furfural (Aa_Fur) were revealed during mixed glucose and xylose fermentation. Carbohydrate metabolism pathways were significantly enriched in response to Aa, while pathways of xenobiotic biodegradation and metabolism were significantly enriched in response to Fur. In addition to these pathways, other pathways were activated in response to Aa_Fur, i.e., cofactor and vitamin metabolism and lipid metabolism. Overexpression of Haa1p or Tye7p improved xylose consumption rates by nearly 50%, while the ethanol yield was enhanced by nearly 8% under acetic acid and furfural stress conditions. Co-overexpression of Haa1p and Tye7p resulted in a 59% increase in xylose consumption rate and a 12% increase in ethanol yield, revealing the beneficial effects of Haa1p and Tye7p on improving the tolerance of yeast to mixed acetic acid and furfural. IMPORTANCE Inhibitor tolerance is essential for S. cerevisiae when fermenting lignocellulosic hydrolysate with various inhibitors, including weak acids, furans, and phenols. The details regarding how xylose-fermenting S. cerevisiae strains respond to multiple inhibitors during fermenting mixed glucose and xylose are still unknown. This study revealed the transcriptional regulation mechanism of an industrial xylose-fermenting S. cerevisiae strain in response to acetic acid and furfural. The transcription factor Haa1p was found to be involved in both acetic acid and furfural tolerance. In addition to Haa1p, four other transcription factors, Hap4p, Yox1p, Tye7p, and Mga1p, were identified as able to improve the resistance of yeast to these two inhibitors. This study underscores the feasibility of uncovering effective transcription factors for constructing robust strains for lignocellulosic bioethanol production.

scholarly journals An Innovative Biocatalyst for Continuous 2G Ethanol Production from Xylo-Oligomers by Saccharomyces cerevisiae through Simultaneous Hydrolysis, Isomerization, and Fermentation (SHIF)

Catalysts ◽  
2019 ◽  
Vol 9 (3) ◽  
pp. 225 ◽  
Author(s):  
Thais Milessi-Esteves ◽  
Felipe Corradini ◽  
Willian Kopp ◽  
Teresa Zangirolami ◽  
Paulo Tardioli ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Production ◽  
Ethanol Yield ◽  
Xylose Isomerase ◽  
Ethanol Productivity ◽  
Spherical Particles ◽  
Birchwood Xylan ◽  
Alginate Gel ◽  
Industrial Implementation ◽  
Ca Alginate

Many approaches have been considered aimed at ethanol production from the hemicellulosic fraction of biomass. However, the industrial implementation of this process has been hindered by some bottlenecks, one of the most important being the ease of contamination of the bioreactor by bacteria that metabolize xylose. This work focuses on overcoming this problem through the fermentation of xylulose (the xylose isomer) by native Saccharomyces cerevisiae using xylo-oligomers as substrate. A new concept of biocatalyst is proposed, containing xylanases and xylose isomerase (XI) covalently immobilized on chitosan, and co-encapsulated with industrial baker’s yeast in Ca-alginate gel spherical particles. Xylo-oligomers are hydrolyzed, xylose is isomerized, and finally xylulose is fermented to ethanol, all taking place simultaneously, in a process called simultaneous hydrolysis, isomerization, and fermentation (SHIF). Among several tested xylanases, Multifect CX XL A03139 was selected to compose the biocatalyst bead. Influences of pH, Ca2+, and Mg2+ concentrations on the isomerization step were assessed. Experiments of SHIF using birchwood xylan resulted in an ethanol yield of 0.39 g/g, (76% of the theoretical), selectivity of 3.12 gethanol/gxylitol, and ethanol productivity of 0.26 g/L/h.

scholarly journals Enhancement of Ethanol Fermentation in Saccharomyces cerevisiae Sake Yeast by Disrupting Mitophagy Function

2013 ◽  
Vol 80 (3) ◽  
pp. 1002-1012 ◽  
Author(s):  
Shodai Shiroma ◽  
Lahiru Niroshan Jayakody ◽  
Kenta Horie ◽  
Koji Okamoto ◽  
Hiroshi Kitagaki
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Yeast Strain ◽  
Parent Strain ◽  
Ethanol Yield ◽  
Synthetic Medium ◽  
Nutrient Concentrations ◽  
Content Type ◽  
Fermentative Capacity ◽  
Limiting Nutrient ◽  
Biomass Resources

ABSTRACTSaccharomyces cerevisiaesake yeast strain Kyokai no. 7 has one of the highest fermentation rates among brewery yeasts used worldwide; therefore, it is assumed that it is not possible to enhance its fermentation rate. However, in this study, we found that fermentation by sake yeast can be enhanced by inhibiting mitophagy. We observed mitophagy in wild-type sake yeast during the brewing of Ginjo sake, but not when the mitophagy gene (ATG32) was disrupted. During sake brewing, the maximum rate of CO2production and final ethanol concentration generated by theatg32Δ laboratory yeast mutant were 7.50% and 2.12% higher than those of the parent strain, respectively. This mutant exhibited an improved fermentation profile when cultured under limiting nutrient concentrations such as those used during Ginjo sake brewing as well as in minimal synthetic medium. The mutant produced ethanol at a concentration that was 2.76% higher than the parent strain, which has significant implications for industrial bioethanol production. The ethanol yield of theatg32Δ mutant was increased, and its biomass yield was decreased relative to the parent sake yeast strain, indicating that theatg32Δ mutant has acquired a high fermentation capability at the cost of decreasing biomass. Because natural biomass resources often lack sufficient nutrient levels for optimal fermentation, mitophagy may serve as an important target for improving the fermentative capacity of brewery yeasts.

scholarly journals Production of Ethanol from Starch by Respiration-Deficient Recombinant Saccharomyces cerevisiae

2005 ◽  
Vol 71 (10) ◽  
pp. 6443-6445 ◽  
Author(s):  
Ebru Toksoy Öner ◽  
Stephen G. Oliver ◽  
Betül Kırdar
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Fermentation ◽  
Ethanol Yield ◽  
Ethanol Productivity ◽  
Recombinant Saccharomyces Cerevisiae ◽  
Direct Fermentation

ABSTRACT A 100%-respiration-deficient nuclear petite amylolytic Saccharomyces cerevisiae NPB-G strain was generated, and its employment for direct fermentation of starch into ethanol was investigated. In a comparison of ethanol fermentation performances with the parental respiration-sufficient WTPB-G strain, the NPB-G strain showed an increase of ca. 48% in both ethanol yield and ethanol productivity.

Improving the ethanol yield by reducing glycerol formation using cofactor regulation in Saccharomyces cerevisiae

2011 ◽  
Vol 33 (7) ◽  
pp. 1375-1380 ◽  
Author(s):  
Liang Zhang ◽  
Yan Tang ◽  
Zhong-peng Guo ◽  
Zhong-yang Ding ◽  
Gui-yang Shi
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Yield ◽  
Glycerol Formation

scholarly journals Increasing Anaerobic Acetate Consumption and Ethanol Yields in Saccharomyces cerevisiae with NADPH-Specific Alcohol Dehydrogenase

2015 ◽  
Vol 81 (23) ◽  
pp. 8108-8117 ◽  
Author(s):  
Brooks M. Henningsen ◽  
Shuen Hon ◽  
Sean F. Covalla ◽  
Carolina Sonu ◽  
D. Aaron Argyros ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Alcohol Dehydrogenase ◽  
Ethanol Yield ◽  
Pentose Phosphate ◽  
Wild Type ◽  
Bifidobacterium Adolescentis ◽  
Acetaldehyde Dehydrogenase ◽  
Content Type ◽  
Sugar Fermentation ◽  
Ethanol Yields

ABSTRACTSaccharomyces cerevisiaehas recently been engineered to use acetate, a primary inhibitor in lignocellulosic hydrolysates, as a cosubstrate during anaerobic ethanolic fermentation. However, the original metabolic pathway devised to convert acetate to ethanol uses NADH-specific acetylating acetaldehyde dehydrogenase and alcohol dehydrogenase and quickly becomes constrained by limited NADH availability, even when glycerol formation is abolished. We present alcohol dehydrogenase as a novel target for anaerobic redox engineering ofS. cerevisiae. Introduction of an NADPH-specific alcohol dehydrogenase (NADPH-ADH) not only reduces the NADH demand of the acetate-to-ethanol pathway but also allows the cell to effectively exchange NADPH for NADH during sugar fermentation. Unlike NADH, NADPH can be freely generated under anoxic conditions, via the oxidative pentose phosphate pathway. We show that an industrial bioethanol strain engineered with the original pathway (expressing acetylating acetaldehyde dehydrogenase fromBifidobacterium adolescentisand with deletions of glycerol-3-phosphate dehydrogenase genesGPD1andGPD2) consumed 1.9 g liter−1acetate during fermentation of 114 g liter−1glucose. Combined with a decrease in glycerol production from 4.0 to 0.1 g liter−1, this increased the ethanol yield by 4% over that for the wild type. We provide evidence that acetate consumption in this strain is indeed limited by NADH availability. By introducing an NADPH-ADH fromEntamoeba histolyticaand with overexpression ofACS2andZWF1, we increased acetate consumption to 5.3 g liter−1and raised the ethanol yield to 7% above the wild-type level.

scholarly journals Gpd1 and Gpd2 Fine-Tuning for Sustainable Reduction of Glycerol Formation in Saccharomyces cerevisiae

2011 ◽  
Vol 77 (17) ◽  
pp. 5857-5867 ◽  
Author(s):  
Georg Hubmann ◽  
Stephane Guillouet ◽  
Elke Nevoigt
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ethanol Yield ◽  
Current Approach ◽  
Fine Tuning ◽  
Anaerobic Growth ◽  
Deletion Strain ◽  
Wild Type ◽  
Double Deletion ◽  
Intermediate Phenotypes ◽  
Glycerol Formation

ABSTRACTGpd1 and Gpd2 are the two isoforms of glycerol 3-phosphate dehydrogenase (GPDH), which is the rate-controlling enzyme of glycerol formation inSaccharomyces cerevisiae.The two isoenzymes play crucial roles in osmoregulation and redox balancing. Past approaches to increase ethanol yield at the cost of reduced glycerol yield have most often been based on deletion of either one or two isogenes (GPD1andGPD2). While single deletions ofGPD1orGPD2reduced glycerol formation only slightly, thegpd1Δgpd2Δ double deletion strain produced zero glycerol but showed an osmosensitive phenotype and abolished anaerobic growth. Our current approach has sought to generate “intermediate” phenotypes by reducing both isoenzyme activities without abolishing them. To this end, theGPD1promoter was replaced in agpd2Δ background by two lower-strengthTEF1promoter mutants. In the same manner, the activity of theGPD2promoter was reduced in agpd1Δbackground. The resulting strains were crossed to obtain different combinations of residualGPD1andGPD2expression levels. Among our engineered strains we identified four candidates showing improved ethanol yields compared to the wild type. In contrast to agpd1Δgpd2Δ double-deletion strain, these strains were able to completely ferment the sugars under quasi-anaerobic conditions in both minimal medium and during simultaneous saccharification and fermentation (SSF) of liquefied wheat mash (wheat liquefact). This result implies that our strains can tolerate the ethanol concentration at the end of the wheat liquefact SSF (up to 90 g liter−1). Moreover, a few of these strains showed no significant reduction in osmotic stress tolerance compared to the wild type.

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