Evolutionary engineering strategies to enhance tolerance of xylose utilizing recombinant yeast to inhibitors derived from spruce biomass
Open accessAbstract
Background: One of the crucial factors for a sustainable and economical production of lignocellulosic based bioethanol is the availability of a robust fermenting microorganism with high tolerance to inhibitors generated during the pretreatment of lignocellulosic raw materials, since these inhibitors are known to severely hinder growth and fermentation. Results: A long-term adaptation in repetitive batch cultures in shake flasks using a cocktail of 12 different inhibitors and a long-term chemostat adaptation using spruce hydrolysate were used as evolutionary engineering strategies to improve the inhibitor tolerance in the metabolically engineered xylose utilizing Saccharomyces cerevisiae strain, TMB3400. The yeast was evolved for a period of 429 and 97 generations in repetitive batch cultures and chemostat cultivation, respectively. During the evolutionary engineering in repetitive batch cultures the maximum specific growth rate increased from 0.18 h-1 to 0.33 h-1 and the time of lag phase was decreased from 48 h to 24 h. In the chemostat adaptation, after 97 generations, the specific conversion rates of HMF and furfural were found to be 3.5 and 4 folds higher respectively, compared to rates after three generations. Two evolved strains (RK60-5, RKU90-3) and one evolved strain (KE1-17) were isolated from evolutionary engineering in repetitive batches and chemostat cultivation, respectively. The strains displayed significantly improved growth performance over TMB3400 when cultivated in spruce hydrolysate under anaerobic conditions, the evolved strains exhibited 25 to 38% increase in specific consumption rate of sugars and 32 to 50% increased specific ethanol productivity compared to TMB3400. The evolved strains RK60-5 and RKU90-3 were unable to consume xylose under anaerobic conditions, whereas, KE1-17 was found to consume xylose at similar rates as TMB3400. Conclusion: Using evolutionary engineering strategies in batch and chemostat cultivations we have generated three evolved strains that show significantly better tolerance to inhibitors in spruce hydrolysate and displayed a shorter time for overall fermentation of sugars compared to the parental strain.
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During ALE, clonal interference, bet hedging, genetic hitchhiking and fluctuating growth environments can lead to significant population heterogeneity.
“Stable phenotypes will accumulate rapidly, although clonal interference [42], bet hedging [43,60], genetic hitchhiking [61] and fluctuating growth environments can lead to significant population heterogeneity [22,62,63].”
From Adaptive laboratory evolution—Principles and applications for biotechnology (2013) · cites this paper as [42]
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- Authors
- Koppram, R., Albers, E., & Olsson, L.
- Affiliations
- Chalmers University of Technology (SE)
- ORCID
- Albers, Eva
- Venue
- Biotechnology for Biofuels
- Published
- 2012-05-11
- Volume
- 5
- Issue
- 1
- Pages
- 32-32
- ISSN
- 1754-6834 (ISSN-L)
- PMC
- PMC3408370
- OpenAlex
- W2131843496
- Cited by
- 160
- References
- 39
- Citations by year
- 2026: 1 · 2025: 6 · 2024: 6 · 2023: 6 · 2022: 13 · 2021: 13 · 2020: 10 · 2019: 10 · 2018: 9 · 2017: 17 · 2016: 20 · 2015: 8 · 2014: 18 · 2013: 19 · 2012: 4
- FWCI
- 8.94
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- cc-by
- Locations
- Biotechnology for Biofuels (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Chalmers Publication Library (Chalmers University of Technology) (submittedVersion)
- Chalmers Research (Chalmers University of Technology) (submittedVersion)
- Funders
- Chalmers Tekniska Högskola
- Energimyndigheten
- Topics
- Physical Sciences › Engineering › Biomedical Engineering › Biofuel production and bioconversion
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
- Keywords
- xylose fermentation, inhibitor tolerance, Saccharomyces cerevisiae, evolutionary engineering, cellulosic ethanol, furfural, 5-hydroxymethylfurfural, ethanol productivity, anaerobic fermentation, recombinant yeast
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