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A next-generation yeast chassis: comprehensive characterization of a robust fast-growing Saccharomyces cerevisiae

ABSTRACT:

Robust and efficient hosts are necessary to facilitate advances in biotechnology. We unveil Saccharomyces cerevisiae strain XP as a promising new microbial chassis owing to its high growth rate and robustness. This strain has a doubling time of 48.2 min, which is 12.8 min faster than the model yeast S. cerevisiae S288C. Strain XP can grow well at low pH (≤ 3.0) and 30% (w/v) glucose under oligotrophic conditions. We establish a genetic manipulation toolbox using URA3 auxotroph and haploid strains, a Cre/loxP-based knocking-out method, and CRISPR/Cas9 genome editing tools. Genomic, transcriptomic and metabolomic data were analyzed in an integrated manner to characterize the strain. The multi-omics analysis reveals that changes in energy and essential nutrient metabolism are linked to rapid growth and robustness. A high-producing strain, XH352-D, was successfully constructed, which produced L-lactic acid with a titer of 52.6 g/L. Here, we comprehensively characterized a robust fast-growing yeast, laying foundation for its potential in industrial production of bulk chemicals.

INSTRUMENT(S): Liquid Chromatography MS - negative - reverse phase, Liquid Chromatography MS - positive - reverse phase

PROVIDER: MTBLS7328 | MetaboLights | 2024-09-26

REPOSITORIES: MetaboLights

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A robust yeast chassis: comprehensive characterization of a fast-growing Saccharomyces cerevisiae.

Long Yangdanyu Y Han Xiao X Meng Xuanlin X Xu Ping P Tao Fei F

mBio 20240112 2

Robust chassis are critical to facilitate advances in synthetic biology. This study describes a comprehensive characterization of a new yeast isolate Saccharomyces cerevisiae XP that grows faster than commonly used research and industrial S. cerevisiae strains. The genomic, transcriptomic, and metabolomic analyses suggest that the fast growth rate is, in part, due to the efficient electron transport chain and key growth factor synthesis. A toolbox for genetic manipulation of the ye ...[more]

PMID: 38214535

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Project description:Saccharomyces cerevisiae is an established microbial host for the production of non-native compounds. The synthesis of these compounds typically demands energy and competes with growth for carbon and energy substrate. Uncoupling product formation form growth would benefit product yields and decrease formation of by-product biomass. Studying non-growing metabolically-active yeast cultures provides a first step towards developing S. cerevisiae as a non-growing, robust cell factory. Non-growing metabolically-active cultures can be obtained in retentostat, a glucose-limited, continuous bioreactor system in which biomass accumulates while spent medium is constantly removed. Hitherto retentostat cultures of S. cerevisiae have only been reported under anaerobiosis, condition inappropriate for the production of energy-demanding products. The present study, using retentostat cultures, explores the physiology of non-dividing, fully respiring S. cerevisiae, focusing on industrially-relevant features. Following model-aided experimental design, retentostat cultivations were optimized for accelerated but smooth transition of S. cerevisiae from exponential growth to near-zero growth rates. During 20 days in retentostat the biomass concentration increased, leading very slow growth rates (specific growth rates below 0.001 h-1) but high culture viability (over 80% of viable cells). The maintenance requirement (mATP) was estimated at 0.64 mmolATP.gX-1.h-1, which is remarkably ca. 35% lower than the mATP measured in anaerobic retentostat cultures. Transcriptional down-regulation of genes involved in biosynthesis and up-regulation of stress-responsive genes towards near-zero growth rates corresponded well with data from anaerobic retentostats. More striking was the extreme heat-shock tolerance of S. cerevisiae, which exceeded by far previously reported heat shock tolerance of notoriously robust yeast cultures such as stationary phase cultures. Furthermore, while the metabolic fluxes in the retentostats were relatively low as a result of extreme caloric restriction, off-line measurements revealed that S. cerevisiae retained a high catabolic capacity. The high viability and extreme heat-shock tolerance revealed the robustness of S. cerevisiae at near-zero growth in retentostat. In addition, the relatively low maintenance requirements and high metabolic capacity under severe calorie restriction underline the potential of S. cerevisiae as a non-dividing microbial cell factory for the production of energy-intensive compounds. The retentostat is a promising tool to identify the molecular basis of this extreme robustness.

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Dataset Information

A next-generation yeast chassis: comprehensive characterization of a robust fast-growing Saccharomyces cerevisiae

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Publications

A robust yeast chassis: comprehensive characterization of a fast-growing <i>Saccharomyces cerevisiae</i>.

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