Industrial Systems Biology of Saccharomyces cerevisiae Enables Novel Succinic Acid Cell Factory
Journal article, 2013

Saccharomyces cerevisiae is the most well characterized eukaryote, the preferred microbial cell factory for the largest industrial biotechnology product (bioethanol), and a robust commerically compatible scaffold to be exploitted for diverse chemical production. Succinic acid is a highly sought after added-value chemical for which there is no native pre-disposition for production and accmulation in S. cerevisiae. The genome-scale metabolic network reconstruction of S. cerevisiae enabled in silico gene deletion predictions using an evolutionary programming method to couple biomass and succinate production. Glycine and serine, both essential amino acids required for biomass formation, are formed from both glycolytic and TCA cycle intermediates. Succinate formation results from the isocitrate lyase catalyzed conversion of isocitrate, and from the alpha-keto-glutarate dehydrogenase catalyzed conversion of alpha-keto-glutarate. Succinate is subsequently depleted by the succinate dehydrogenase complex. The metabolic engineering strategy identified included deletion of the primary succinate consuming reaction, Sdh3p, and interruption of glycolysis derived serine by deletion of 3-phosphoglycerate dehydrogenase, Ser3p/Ser33p. Pursuing these targets, a multi-gene deletion strain was constructed, and directed evolution with selection used to identify a succinate producing mutant. Physiological characterization coupled with integrated data analysis of transcriptome data in the metabolically engineered strain were used to identify 2nd-round metabolic engineering targets. The resulting strain represents a 30-fold improvement in succinate titer, and a 43-fold improvement in succinate yield on biomass, with only a 2.8-fold decrease in the specific growth rate compared to the reference strain. Intuitive genetic targets for either over-expression or interruption of succinate producing or consuming pathways, respectively, do not lead to increased succinate. Rather, we demonstrate how systems biology tools coupled with directed evolution and selection allows non-intuitive, rapid and substantial re-direction of carbon fluxes in S. cerevisiae, and hence show proof of concept that this is a potentially attractive cell factory for over-producing different platform chemicals.

strains

growth

scale metabolic model

validation

in-silico

yeast

reconstruction

bioethanol

products

biotechnology

Author

José Manuel Otero

Chalmers, Chemical and Biological Engineering, Life Sciences

D. Cimini

University of Campania Luigi Vanvitelli

K. R. Patil

European Molecular Biology Laboratory

Technical University of Denmark (DTU)

S. G. Poulsen

Technical University of Denmark (DTU)

Lisbeth Olsson

Chalmers, Chemical and Biological Engineering, Industrial biotechnology

Jens B Nielsen

Chalmers, Chemical and Biological Engineering, Life Sciences

PLoS ONE

1932-6203 (ISSN) 19326203 (eISSN)

Vol. 8 1 e54144

Driving Forces

Sustainable development

Areas of Advance

Energy

Life Science Engineering (2010-2018)

Subject Categories

Chemical Sciences

DOI

10.1371/journal.pone.0054144

More information

Latest update

5/9/2018 2