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.