Abstract
Genetically customised Saccharomyces cerevisiae that can produce ethanol and additional bio-based chemicals from sustainable agro-industrial feedstocks (for example, residual plant biomass) are of major interest to the biofuel industry. We investigated the microbial biorefinery concept of ethanol and squalene co-production using S. cerevisiae (strain YUG37-ERG1) wherein ERG1 (squalene epoxidase) transcription is under the control of a doxycycline-repressible tet0 7 -CYC1 promoter. The production of ethanol and squalene by YUG37-ERG1 grown using agriculturally sourced grass juice supplemented with doxycycline was assessed. Use of the tet0 7 -CYC1 promoter permitted regulation of ERG1 expression and squalene accumulation in YUG37-ERG1, allowing us to circumvent the lethal growth phenotype seen when ERG1 is disrupted completely. In experiments using grass juice feedstock supplemented with 0 to 50 μg doxycycline mL-1, YUG37-ERG1 fermented ethanol (22.5 [±0.5] mg mL-1) and accumulated the highest squalene content (7.89 ± 0.25 mg g-1 dry biomass) and yield (18.0 ± 4.18 mg squalene L-1) with supplements of 5.0 and 0.025 μg doxycycline mL-1, respectively. Grass juice was found to be rich in water-soluble carbohydrates (61.1 [±3.6] mg sugars mL-1) and provided excellent feedstock for growth and fermentation studies using YUG37-ERG1. Residual plant biomass components from crop production and rotation systems represent possible substrates for microbial fermentation of biofuels and bio-based compounds. This study is the first to utilise S. cerevisiae for the co-production of ethanol and squalene from grass juice. Our findings underscore the value of the biorefinery approach and demonstrate the potential to integrate microbial bioprocess engineering with existing agriculture.
Highlights
Customised Saccharomyces cerevisiae that can produce ethanol and additional bio-based chemicals from sustainable agro-industrial feedstocks are of major interest to the biofuel industry
We investigated the potential to produce squalene as a bio-based chemical product of yeast fermentation using a customised S. cerevisiae strain (YUG37-squalene epoxidase (ERG1)) wherein ERG1 gene transcription is under the control of a doxycycline-repressible promoter that replaces the ERG1 promoter at the chromosomal locus [35,36]
Of the many microorganisms that could be employed for ethanol production, S. cerevisiae remains the species of choice in industrial-scale fermentation processes [45]
Summary
Customised Saccharomyces cerevisiae that can produce ethanol and additional bio-based chemicals from sustainable agro-industrial feedstocks (for example, residual plant biomass) are of major interest to the biofuel industry. We investigated the microbial biorefinery concept of ethanol and squalene co-production using S. cerevisiae (strain YUG37-ERG1) wherein ERG1 (squalene epoxidase) transcription is under the control of a doxycycline-repressible tet07-CYC1 promoter. Microorganisms that possess the enzymatic machinery needed to unlock fuel energy from cellulosic and lignocellulosic fractions of plant biomass [1,2,3,4,5] and recombinant strains that can utilise alternative substrates (such as inulin [6]) for the production of additional bio-based products [7,8,9,10], are of major interest to biofuel and biorefinery industries. Given the increasing commercial demand for squalene alongside growing international concern for the fate of food crops and the exploitation of marine habitats, sustainable sources of squalene are required
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