Combined engineering of disaccharide transport and phosphorolysis for enhanced ATP yield from sucrose fermentation in Saccharomyces cerevisiae

Author(s):Show less -

Marques, Wesley Leoricy ^{[1, 2, 3]} ; Mans, Robert ^[2] ; Henderson, Ryan K. ^{[4, 5]} ; Marella, Eko Roy ^[2] ; ter Horst, Jolanda ^[2] ; de Hulster, Erik ^[2] ; Poolman, Bert ^{[4, 5]} ; Daran, Jean-Marc ^[2] ; Pronk, Jack T. ^[2] ; Gombert, Andreas K. ^[1] ; van Maris, Antonius J. A. ^{[2, 6]}

Affiliation

^[1] Univ Estadual Campinas, Sch Food Engn, Rua Monteiro Lobato 80, BR-13083862 Campinas, SP - Brazil

^[2] Delft Univ Technol, Dept Biotechnol, Van der Maasweg 9, NL-2629 HZ Delft - Netherlands

^[3] Univ Sao Paulo, Dept Chem Engn, Ave Prof Lineu Prestes 580, Bloco 20, BR-05424970 Sao Paulo, SP - Brazil

^[4] Univ Groningen, Zernike Inst Adv Mat, Nijenborgh 4, NL-9747 AG Groningen - Netherlands

^[5] Univ Groningen, Dept Biochem, Groningen Biomol Sci & Biotechnol Inst, Nijenborgh 4, NL-9747 AG Groningen - Netherlands

^[6] AlbaNova Univ Ctr, KTH Royal Inst Technol, Sch Biotechnol, Div Ind Biotechnol, SE-10691 Stockholm - Sweden

• Document type: Journal article
• Source: METABOLIC ENGINEERING; v. 45, p. 121-133, JAN 2018.
• Web of Science Citations: 8

Abstract

Anaerobic industrial fermentation processes do not require aeration and intensive mixing and the accompanying cost savings are beneficial for production of chemicals and fuels. However, the free-energy conservation of fermentative pathways is often insufficient for the production and export of the desired compounds and/or for cellular growth and maintenance. To increase free-energy conservation during fermentation of the industrially relevant disaccharide sucrose by Saccharomyces cerevisiae, we first replaced the native yeast alpha-glucosidases by an intracellular sucrose phosphorylase from Leuconostoc mesenteroides (LmSPase). Subsequently, we replaced the native proton-coupled sucrose uptake system by a putative sucrose facilitator from Phaseolus vulgaris (PvSUF1). The resulting strains grew anaerobically on sucrose at specific growth rates of 0.09 +/- 0.02 h(-1) (LmSPase) and 0.06 +/- 0.01 h(-1) (PvSUF1, LmSPase). Overexpression of the yeast PGM2 gene, which encodes phosphoglucomutase, increased anaerobic growth rates on sucrose of these strains to 0.23 +/- 0.01 h(-1) and 0.08 +/- 0.00 h(-1), respectively. Determination of the biomass yield in anaerobic sucrose-limited chemostat cultures was used to assess the free-energy conservation of the engineered strains. Replacement of intracellular hydrolase with a phosphorylase increased the biomass yield on sucrose by 31%. Additional replacement of the native proton-coupled sucrose uptake system by PvSUF1 increased the anaerobic biomass yield by a further 8%, resulting in an overall increase of 41%. By experimentally demonstrating an energetic benefit of the combined engineering of disaccharide uptake and cleavage, this study represents a first step towards anaerobic production of compounds whose metabolic pathways currently do not conserve sufficient free-energy. (AU)

• FAPESP's process: 12/05548-1 - Metabolic engineering of Saccharomyces cerevisiae aimed at improving the energetic yield of sucrose metabolism
• Grantee: Wesley Leoricy Marques
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 12/16630-0 - Metabolic engineering of Saccharomyces cerevisiae aimed at improving the energetic yield of sucrose metabolism
• Grantee: Wesley Leoricy Marques
• Support Opportunities: Scholarships abroad - Research Internship - Master's degree