Engineering Pichia pastoris for improved NADH regeneration: A novel chassis strain for whole-cell catalysis

• Martina Geier,
• Christoph Brandner,
• Gernot A. Strohmeier,
• Mélanie Hall,
• Franz S. Hartner and
• Anton Glieder

Beilstein J. Org. Chem. 2015, 11, 1741–1748, doi:10.3762/bjoc.11.190

• yeast P. pastoris by engineering its methanol utilization pathway. By deleting the pathway for methanol assimilation, we forced flux through the dissimilatory pathway in which theoretically two molecules of NADH can be formed per molecule of methanol. The resulting double knock-out strain P. pastoris

• )-acetoin: 2.32 min, (2R,3R)-butane-2,3-diol: 8.27 min, meso-butane-2,3-diol: 8.90 min. Simplified schematic representation of the methanol utilization pathway in Pichia pastoris. The main pathways and the respective enzymes are shown. AOX: alcohol oxidase; FLD: glutathione-dependent formaldehyde

• utilization pathway of Pichia pastoris for improved NADH regeneration. By deleting the genes coding for dihydroxyacetone synthase isoform 1 and 2 (DAS1 and DAS2), NADH regeneration via methanol oxidation (dissimilation) was increased significantly. The resulting Δdas1 Δdas2 strain performed better in