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Title: Domesticating a food spoilage yeast into an organic acid–tolerant metabolic engineering host: Lactic acid production by engineered Zygosaccharomyces bailii

Journal Article · · Biotechnology and Bioengineering
DOI:https://doi.org/10.1002/bit.27576· OSTI ID:1764380
 [1];  [1];  [2]; ORCiD logo [1]
  1. Univ. of Illinois at Urbana-Champaign, IL (United States)
  2. Univ. of Illinois at Urbana-Champaign, IL (United States); USDA‐ARS Peoria, IL (United States)
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Lactic acid represents an important class of commodity chemicals, which can be produced by microbial cell factories. However, due to the toxicity of lactic acid at lower pH, microbial production requires the usage of neutralizing agents to maintain neutral pH. Zygosaccharomyces bailii, a food spoilage yeast, can grow under the presence of organic acids used as food preservatives. This unique trait of the yeast might be useful for producing lactic acid. With the goal of domesticating the organic acid-tolerant yeast as a metabolic engineering host, seven Z. bailii strains were screened in a minimal medium with 10 g/L of acetic, or 60 g/L of lactic acid at pH 3. The Z. bailii NRRL Y7239 strain was selected as the most robust strain to be engineered for lactic acid production. By applying a PAN-ARS-based CRISPR-Cas9 system consisting of a transfer RNA promoter and NAT selection, we demonstrated the targeted deletion of ADE2 and site-specific integration of Rhizopus oryzae ldhA coding for lactate dehydrogenase into the PDC1 locus. The resulting pdc1::ldhA strain produced 35 g/L of lactic acid without ethanol production. This study demonstrates the feasibility of the CRISPR-Cas9 system in Z. bailii, which can be applied for a fundamental study of the species.

Research Organization:
Center for Advanced Bioenergy and Bioproducts Innovation (CABBI), Urbana, IL (United States)
Sponsoring Organization:
USDOE Office of Science (SC), Biological and Environmental Research (BER)
Grant/Contract Number:
SC0018420
OSTI ID:
1764380
Alternate ID(s):
OSTI ID: 1786089; OSTI ID: 1991873
Journal Information:
Biotechnology and Bioengineering, Vol. 118, Issue 1; ISSN 0006-3592
Publisher:
WileyCopyright Statement
Country of Publication:
United States
Language:
English

References (42)

Pyruvate decarboxylase: An indispensable enzyme for growth of Saccharomyces cerevisiae on glucose journal March 1996
Advances in molecular tools for the use of Zygosaccharomyces bailii as host for biotechnological productions and construction of  the first auxotrophic mutant: Improved molecular tools for Z. bailii manipulation journal July 2010
Extreme resistance to weak-acid preservatives in the spoilage yeast Zygosaccharomyces bailii journal August 2013
A gRNA-tRNA array for CRISPR-Cas9 based rapid multiplexed genome editing in Saccharomyces cerevisiae journal March 2019
A comparative study of ethanol production by Issatchenkia orientalis strains under stress conditions journal January 2012
Looking beyond Saccharomyces : the potential of non-conventional yeast species for desirable traits in bioethanol fermentation journal June 2015
Lactic acid production from xylose by engineered Saccharomyces cerevisiae without PDC or ADH deletion journal June 2015
Controlled mixed culture fermentation: a new perspective on the use of non- Saccharomyces   yeasts in winemaking journal March 2010
l-Lactic acid production from glucose and xylose with engineered strains of Saccharomyces cerevisiae: aeration and carbon source influence yields and productivities journal April 2018
The ADE2 gene from Saccharomyces cerevisiae: sequence and new vectors journal October 1990
Sphingolipids contribute to acetic acid resistance in Zygosaccharomyces bailii: Sphingolipids Contribute to Acetic Acid Resistance journal December 2015
Growth requirements of pyruvate-decarboxylase-negative Saccharomyces cerevisiae journal May 1999
Assessing physio-macromolecular effects of lactic acid on Zygosaccharomyces bailii cells during microaerobic fermentation journal July 2016
Enzymology of the pigmented adenine-requiring mutants of Saccharomyces and Schizosaccharomyces journal February 1969
16 years research on lactic acid production with yeast – ready for the market? journal January 2010
Lactic acid properties, applications and production: A review journal March 2013
Abc1p Is a Multidrug Efflux Transporter That Tips the Balance in Favor of Innate Azole Resistance in Candida krusei journal November 2008
Impurity and cost considerations for nutrient supplementation of whey permeate fermentations to produce lactic acid for biodegradable plastics journal January 2003
An autonomously replicating sequence for use in a wide range of budding yeasts journal December 2013
Modification of metabolic pathways of Saccharomyces cerevisiae by the expression of lactate dehydrogenase and deletion of pyruvate decarboxylase genes for the lactic acid fermentation at low pH value journal January 1998
High-efficiency CRISPR/Cas9 multiplex gene editing using the glycine tRNA-processing system-based strategy in maize journal August 2016
sgRNAcas9: A Software Package for Designing CRISPR sgRNA and Evaluating Potential Off-Target Cleavage Sites journal June 2014
Development of a CRISPR/Cas9 system for high efficiency multiplexed gene deletion in Rhodosporidium toruloides journal May 2019
Transcriptional Response to Lactic Acid Stress in the Hybrid Yeast Zygosaccharomyces parabailii journal December 2017
Evolutionary restoration of fertility in an interspecies hybrid yeast, by whole-genome duplication after a failed mating-type switch journal May 2017
Genome editing in Kluyveromyces and Ogataea yeasts using a broad-host-range Cas9/gRNA co-expression plasmid journal February 2018
CRISPR-Cas9 mediated gene deletions in lager yeast Saccharomyces pastorianus journal December 2017
Improvement of L -Lactate Production by CYB2 Gene Disruption in a Recombinant Saccharomyces cerevisiae Strain under Low pH Condition journal November 2008
Proposal of Zygosaccharomyces parabailii sp. nov. and Zygosaccharomyces pseudobailii sp. nov., novel species closely related to Zygosaccharomyces bailii journal May 2013
The Fate of Acetic Acid during Glucose Co-Metabolism by the Spoilage Yeast Zygosaccharomyces bailii journal December 2012
The expanding RNA polymerase III transcriptome journal December 2007
The spoilage yeast Zygosaccharomyces bailii : Foe or friend?: The spoilage yeast Zygosaccharomyces bailii : foe or friend? journal June 2017
The Effect of Pyruvate Decarboxylase Gene Knockout in Saccharomyces cerevisiae on L -Lactic Acid Production journal May 2006
Transcription by RNA polymerase III: more complex than we thought journal May 2011
Population genomics shows no distinction between pathogenic Candida krusei and environmental Pichia kudriavzevii: One species, four names journal July 2018
CRISPR-mediated genome editing in non-conventional yeasts for biotechnological applications journal April 2019
Metabolic Engineering of Saccharomyces cerevisiae for Efficient Production of Pure L-(+)-Lactic Acid journal January 2006
Effect of benzoic acid on metabolic fluxes in yeasts: A continuous-culture study on the regulation of respiration and alcoholic fermentation journal July 1992
Engineering Kluyveromyces marxianus as a Robust Synthetic Biology Platform Host journal September 2018
Low‐pH production of d ‐lactic acid using newly isolated acid tolerant yeast Pichia kudriavzevii NG7 journal June 2018
Isolation of the DLD gene of Saccharomyces cerevisiae encoding the mitochondrial enzyme D-lactate ferricytochrome c oxidoreductase journal April 1993
Boosting CRISPR/Cas9 multiplex editing capability with the endogenous tRNA-processing system journal March 2015

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59 BASIC BIOLOGICAL SCIENCES
CRISPR Cas9
Zygosaccharomyces bailii
lactic acid
metabolic engineering