Resistance of yeast to inhibitors of lignocellulosic bioethanol production

Moreno-Martínez, Elena (2019) Resistance of yeast to inhibitors of lignocellulosic bioethanol production. PhD thesis, University of Nottingham.

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As we witness the depletion of natural fossil fuel resources, there is an increasing urgency to find and develop sustainable alternatives that can still meet the world’s ever growing needs. Lignocellulose is one of the most abundant sources for biomass-based energy, a promising candidate for a potentially powerful new combustible. Considerable effort has been made to obtain biofuel from lignocellulosic materials using microorganisms, but there are still a number of constraints to be overcome for this new sustainable form of energy to become the ultimate solution to the petroleum problem. Upon breakdown of lignocellulosic biomass to release the sugars that will be used by the ethanologenic organisms, numerous toxic compounds are also generated; weak acids, furans and phenols impair microbial growth and fermentation, thus reducing efficiency of the process and increasing the cost.

The first part of this project focuses on improving resistance in the ethanologenic model organism Saccharomyces cerevisiae against the strongly inhibitory compounds furfural, acetic acid and benzoic acid. For this purpose, a novel RNAi-based evolution approach was used to obtain strains with accumulated silenced genes that together confer increased resistance to these individual inhibitors. RNAi here provided a useful tool for accelerated evolution of strains towards increased resistance to these stressors, especially acetic acid, and enabled the discovery of multiple genes whose silencing or deletion conferred increased resistance to one or multiple inhibitors. This strategy uncovered previously unknown synergistic interactions between two or more genes that contribute to resistance to acetic acid, combinatorial effects that could not have been predicted or obtained through other approaches.

The rest of this work digs into another layer concerning resistance to lignocellulosic-derived inhibitors; the variability in resistance across individual cells within a clonal yeast population, also called phenotypic heteroresistance. Understanding individual-cell effects of hydrolysate inhibitors could help engineering of population-level resistance. The degree of heteroresistance was specific to vanillin, furfural, benzoic and acetic acids in the fermenting yeast S. cerevisiae. Inclusion of a second inhibitor altered heteroresistance relative to either inhibitor alone, with two main patterns observed for the tested combinations. Links between adaptive resistance and adaptive heterogeneity after short pretreatments with subinhibitory concentrations of the inhibitors were also investigated. Acclimation to acetic and benzoic acids triggered a quick short-term adaptive response, with different changes in heteroresistance associated, for which cytosolic pH might be partially responsible.

Findings derived from this research provide better understanding of the behaviour of yeast in lignocellulosic ethanol fermentations, as well as multiple potential targets for strain optimisation to help improve production.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: Avery, Simon
Powell, Chris
Keywords: Bioethanol; Yeast; Ligenocellulosic hydrolysates; Phenotypic heterogeneity
Subjects: T Technology > TP Chemical technology
Faculties/Schools: UK Campuses > Faculty of Medicine and Health Sciences > School of Life Sciences
Item ID: 55927
Depositing User: Moreno Martínez, Elena
Date Deposited: 19 Jul 2019 04:40
Last Modified: 19 Jul 2021 04:30

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