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Transcriptomic analysis of synchrony and productivity in self-cycling fermentation of engineered yeast producing shikimic acid.

ABSTRACT: Industrial fermentation provides a wide variety of bioproducts, such as food, biofuels and pharmaceuticals. Self-cycling fermentation (SCF), an advanced automated semi-continuous fermentation approach, has shown significant advantages over batch reactors (BR); including cell synchrony and improved production. Here, Saccharomyces cerevisiae engineered to overproduce shikimic acid was grown under SCF operation. This led to four-fold increases in product yield and volumetric productivity compared to BR. Transcriptomic analyses were performed to understand the cellular mechanisms leading to these increases. Results indicate an up-regulation of a large number of genes related to the cell cycle and DNA replication in the early stages of SCF cycles, inferring substantial synchronization. Moreover, numerous genes related to gluconeogenesis, the citrate cycle and oxidative phosphorylation were significantly up-regulated in the late stages of SCF cycles, consistent with significant increases in shikimic acid yield and productivity.

SUBMITTER: Tan Y

PROVIDER: S-EPMC8660916 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature

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Transcriptomic analysis of synchrony and productivity in self-cycling fermentation of engineered yeast producing shikimic acid.

Tan Yusheng Y Agustin Roman Vincent C RVC Stein Lisa Y LY Sauvageau Dominic D

Biotechnology reports (Amsterdam, Netherlands) 20211203

Industrial fermentation provides a wide variety of bioproducts, such as food, biofuels and pharmaceuticals. Self-cycling fermentation (SCF), an advanced automated semi-continuous fermentation approach, has shown significant advantages over batch reactors (BR); including cell synchrony and improved production. Here, <i>Saccharomyces cerevisiae</i> engineered to overproduce shikimic acid was grown under SCF operation. This led to four-fold increases in product yield and volumetric productivity com ...[more]

PMID: 34934640

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Project description:The cellulosic ethanol industry has developed efficient strategies for converting sugars obtained from various cellulosic feedstocks to bioethanol. However, any further major improvements in ethanol productivity will require development of novel and innovative fermentation strategies that enhance incumbent technologies in a cost-effective manner. The present study investigates the feasibility of applying self-cycling fermentation (SCF) to cellulosic ethanol production to elevate productivity. SCF is a semi-continuous cycling process that employs the following strategy: once the onset of stationary phase is detected, half of the broth volume is automatically harvested and replaced with fresh medium to initiate the next cycle. SCF has been shown to increase product yield and/or productivity in many types of microbial cultivation. To test whether this cycling process could increase productivity during ethanol fermentations, we mimicked the process by manually cycling the fermentation for five cycles in shake flasks, and then compared the results to batch operation.Mimicking SCF for five cycles resulted in regular patterns with regards to glucose consumption, ethanol titer, pH, and biomass production. Compared to batch fermentation, our cycling strategy displayed improved ethanol volumetric productivity (the titer of ethanol produced in a given cycle per corresponding cycle time) and specific productivity (the amount of ethanol produced per cellular biomass) by 43.1 ± 11.6 and 42.7 ± 9.8%, respectively. Five successive cycles contributed to an improvement of overall productivity (the aggregate amount of ethanol produced at the end of a given cycle per total processing time) and the estimated annual ethanol productivity (the amount of ethanol produced per year) by 64.4 ± 3.3 and 33.1 ± 7.2%, respectively.This study provides proof of concept that applying SCF to ethanol production could significantly increase productivities, which will help strengthen the cellulosic ethanol industry.

Project description:BackgroundEfficient production of SA in Escherichia coli has been achieved by modifying key genes of the central carbon metabolism and SA pathway, resulting in overproducing strains grown in batch- or fed-batch-fermentor cultures using a complex broth including glucose and YE. In this study, we performed a GTA to identify those genes significantly upregulated in an engineered E. coli strain, PB12.SA22, in mid EXP (5 h), early STA (STA1, 9 h), and late STA (STA2, 44 h) phases, grown in complex fermentation broth in batch-fermentor cultures.ResultsGrowth of E. coli PB12.SA22 in complex fermentation broth for SA production resulted in an EXP growth during the first 9 h of cultivation depending of supernatant available aromatic amino acids provided by YE because, when tryptophan was totally consumed, cells entered into a second, low-growth phase (even in the presence of glucose) until 26 h of cultivation. At this point, glucose was completely consumed but SA production continued until the end of the fermentation (50 h) achieving the highest accumulation (7.63 g/L of SA). GTA between EXP/STA1, EXP/STA2 and STA1/STA2 comparisons showed no significant differences in the regulation of genes encoding enzymes of central carbon metabolism as in SA pathway, but those genes encoding enzymes involved in sugar, amino acid, nucleotide/nucleoside, iron and sulfur transport; amino acid catabolism and biosynthesis; nucleotide/nucleoside salvage; acid stress response and modification of IM and OM were upregulated between comparisons.ConclusionsGTA during SA production in batch-fermentor cultures of strain PB12.SA22 grown in complex fermentation broth during the EXP, STA1 and STA2 phases was studied. Significantly, upregulated genes during the EXP and STA1 phases were associated with transport, amino acid catabolism, biosynthesis, and nucleotide/nucleoside salvage. In STA2, upregulation of genes encoding transporters and enzymes involved in the synthesis and catabolism of Arg suggests that this amino acid could have a key role in the fuelling of carbon toward SA synthesis, whereas upregulation of genes involved in pH stress response, such as membrane modifications, suggests a possible response to environmental conditions imposed on the cell at the end of the fermentation.

Dataset Information

Transcriptomic analysis of synchrony and productivity in self-cycling fermentation of engineered yeast producing shikimic acid.

Publications

Transcriptomic analysis of synchrony and productivity in self-cycling fermentation of engineered yeast producing shikimic acid.

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