title: Increasing free-energy (ATP) conservation in maltose-grown Saccharomyces cerevisiae by expression of a heterologous maltose phosphorylase.
abstract: Increasing free-energy conservation from the conversion of substrate into product is crucial for further development of many biotechnological processes. In theory, replacing the hydrolysis of disaccharides by a phosphorolytic cleavage reaction provides an opportunity to increase the ATP yield on the disaccharide. To test this concept, we first deleted the native maltose metabolism genes in Saccharomyces cerevisiae. The knockout strain showed no maltose-transport activity and a very low residual maltase activity (0.03 μmol mg protein(-1)min(-1)). Expression of a maltose phosphorylase gene from Lactobacillus sanfranciscensis and the MAL11 maltose-transporter gene resulted in relatively slow growth (μ(aerobic) 0.09 ± 0.03 h(-1)). Co-expression of Lactococcus lactis β-phosphoglucomutase accelerated maltose utilization via this route (μ(aerobic) 0.21 ± 0.01 h(-1), μ(anaerobic) 0.10 ± 0.00 h(-1)). Replacing maltose hydrolysis with phosphorolysis increased the anaerobic biomass yield on maltose in anaerobic maltose-limited chemostat cultures by 26%, thus demonstrating the potential of phosphorolysis to improve the free-energy conservation of disaccharide metabolism in industrial microorganisms.
authors.affiliation: Department of Biotechnology, Delft University of Technology and Kluyver Centre for Genomics of Industrial Fermentation, Julianalaan 67, Delft, The Netherlands.
authors.forename: Stefan
authors.initials: S
authors.lastname: de Kok
authors.forename: Duygu
authors.initials: D
authors.lastname: Yilmaz
authors.forename: Erwin
authors.initials: E
authors.lastname: Suir
authors.forename: Jack T
authors.initials: JT
authors.lastname: Pronk
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Daran
authors.forename: Antonius J A
authors.initials: AJ
authors.lastname: van Maris
mesh_terms: D000255:Adenosine Triphosphate / Q000096:biosynthesis*; D000693:Anaerobiosis / Q000187:drug effects / Q000235:genetics; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D055785:Gene Knockdown Techniques; D005964:Glucosyltransferases / Q000096:biosynthesis / Q000235:genetics; D007778:Lactobacillus / Q000201:enzymology / Q000235:genetics; D008320:Maltose / Q000378:metabolism* / Q000494:pharmacology; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism; D030781:Organisms, Genetically Modified / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D013549:Sweetening Agents / Q000378:metabolism / Q000494:pharmacology; D027981:Symporters / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D009004:Monosaccharide Transport Proteins; D029701:Saccharomyces cerevisiae Proteins; D013549:Sweetening Agents; D027981:Symporters; C067575:maltose transport system, S cerevisiae; D008320:Maltose; D000255:Adenosine Triphosphate; D005964:Glucosyltransferases; C021535:maltose phosphorylase