Towards the exploitation of glycerol's high reducing power in Saccharomyces cerevisiae-based bioprocesses |
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| Institution: | 1. Metabolic and Biomolecular Engineering National Research Laboratory, Department of Chemical and Biomolecular Engineering (BK21 Plus program), BioProcess Engineering Research Center, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea;2. Center for Systems and Synthetic Biotechnology, Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea;1. Department of Food Science and Human Nutrition, Agricultural University of Athens, Iera Odos 75, Athens, Greece;2. Department of Chemical Engineering, Lund University, P.O. Box 124, 221 00 Lund, Sweden;1. Department of Chemical Engineering and Applied Chemistry, University of Toronto, Canada;2. Institute of Biomaterials and Biomedical Engineering (IBBME), University of Toronto, Canada;1. Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, Republic of Korea;2. Department of Food Science and Human Nutrition, and Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA;1. School of Energy and Environment, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong, China;2. Microbial Processes and Interactions, TERRA Teaching and Research Center, University of Liège – Gembloux Agro-Bio Tech, Av. De la Faculté, 2B, 5030 Gembloux, Belgium |
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| Abstract: | One advantage of using glycerol as a carbon source for industrial bioprocesses is its higher degree of reduction compared to glucose. In order to exploit this reducing power for the production of reduced compounds thereby significantly increasing maximum theoretical yields, the electrons derived from glycerol oxidation must first be saved in the form of cytosolic NAD(P)H. However, the industrial platform organism Saccharomyces cerevisiae naturally uses an FAD-dependent pathway for glycerol catabolism transferring the electrons to the respiratory chain. Here, we developed a pathway replacement strategy forcing glycerol catabolism through a synthetic, NAD+-dependent route. The required expression cassettes were integrated via CRISPR-Cas9 targeting the endogenous GUT1 locus, thereby abolishing the native FAD-dependent pathway. Interestingly, this pathway replacement even established growth in synthetic glycerol medium of strains naturally unable to grow on glycerol and an engineered derivative of CEN.PK even showed the highest ever reported maximum specific growth rate on glycerol (0.26 h−1). |
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| Keywords: | Yeast Glycerol catabolism DHA pathway NADH |
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