Enhanced tolerance of Saccharomyces cerevisiae to multiple lignocellulose-derived inhibitors through modulation of spermidine contents.
Fermentation inhibitors present in lignocellulose hydrolysates are inevitable obstacles for achieving economic production of biofuels and biochemicals by industrial microorganisms. Here we show that spermidine (SPD) functions as a chemical elicitor for enhanced tolerance of Saccharomyces cerevisiae against major fermentation inhibitors. In addition, the feasibility of constructing an engineered S. cerevisiae strain capable of tolerating toxic levels of the major inhibitors without exogenous addition of SPD was explored. Specifically, we altered expression levels of the genes in the SPD biosynthetic pathway. Also, OAZ1 coding for ornithine decarboxylase (ODC) antizyme and TPO1 coding for the polyamine transport protein were disrupted to increase intracellular SPD levels through alleviation of feedback inhibition on ODC and prevention of SPD excretion, respectively. Especially, the strain with combination of OAZ1 and TPO1 double disruption and overexpression of SPE3 not only contained spermidine content of 1.1mg SPD/g cell, which was 171% higher than that of the control strain, but also exhibited 60% and 33% shorter lag-phase period than that of the control strain under the medium containing furan derivatives and acetic acid, respectively. While we observed a positive correlation between intracellular SPD contents and tolerance phenotypes among the engineered strains accumulating different amounts of intracellular SPD, too much SPD accumulation is likely to cause metabolic burden. Therefore, genetic perturbations for intracellular SPD levels should be optimized in terms of metabolic burden and SPD contents to construct inhibitor tolerant yeast strains. We also found that the genes involved in purine biosynthesis and cell wall and chromatin stability were related to the enhanced tolerance phenotypes to furfural. The robust strains constructed in this study can be applied for producing chemicals and advanced biofuels from cellulosic hydrolysates.
Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, Republic of Korea.
Sun-Ki
SK
Kim
Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA; Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Yong-Su
YS
Jin
College of Life sciences and Biotechnology, Korea University, Seoul 136-713, Republic of Korea.
In-Geol
IG
Choi
Department of Bio and Fermentation Convergence, Kookmin University, Seoul 136-702, Republic of Korea.
Yong-Cheol
YC
Park
Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, Republic of Korea. Electronic address: jhseo94@snu.ac.kr.
Jin-Ho
JH
Seo
D017920:Antiporters / Q000235:genetics / Q000378:metabolism; D025141:Drug Resistance, Fungal; D008031:Lignin / Q000378:metabolism / Q000494:pharmacology; D060847:Metabolic Engineering; D027701:Organic Cation Transport Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013095:Spermidine / Q000096:biosynthesis*
D017920:Antiporters; C497099:Oaz1 protein, S cerevisiae; D027701:Organic Cation Transport Proteins; D029701:Saccharomyces cerevisiae Proteins; C475058:TPO1 protein, S cerevisiae; C036909:lignocellulose; D008031:Lignin; D013095:Spermidine
Biofuels; Feedback inhibition; Fermentation inhibitors; Polyamine transport protein; RNA sequencing; Spermidine