Engineering of Saccharomyces cerevisiae for the production of l-glycerol 3-phosphate

Abstract

l-glycerol 3-phosphate ( l-G3P) was accumulated in Saccharomyces cerevisiae by pathway engineering. Intracellular concentration of this metabolic intermediate could be increased more than 20 times compared to the wild type by overexpressing GPD1 encoding the glycerol 3-phosphate dehydrogenase in a gpp1Δ gpp2Δ mutant which lacks both isoenzymes of glycerol 3-phosphatase. Investigation of cellular pattern of triacylglycerols and glycerophospholipids did not reveal considerable changes due to accumulation of their precursor l-G3P. Hyperosmotic stress did not affect the l-G3P pool in the gpp1Δ gpp2Δ mutant overexpressing GPD1 despite an about 4-fold increase of specific GPD activity. In contrast, oxygen limitation improved intracellular l-G3P concentration by enhancing the availability of cytosolic NADH. The reduction of pyruvate decarboxylase activity by deleting PDC2 led to an additional increase. In fact, the triple mutant gpp1Δ gpp2Δ pdc2Δ overexpressing GPD1 accumulated 17 mg l-G3P/g dry weight during glucose batch fermentation under oxygen limitation. This value corresponds to an about 100-fold increase compared to that found in the wild type.