%0 Journal Article %T NADPH-Auxotrophic E-coil: A Sensor Strain for Testing in Vivo Regeneration of NADPH %+ Max Planck Institute of Molecular Physiology %+ Génomique métabolique (UMR 8030) %A Lindner, Steffen N. %A Ramirez, Liliana Calzadiaz %A Kruesemann, Jan L. %A Yishai, Oren %A Belkhelfa, Sophia %A He, Hai %A Bouzon, Madeleine %A Doring, Volker %A Bar-Even, Arren %Z Max Planck Society %Z China Scholarship Council %Z Energy Sustainability Grant of the National Council of Science and Technology [429271] %Z Mexican Ministry of Energy (CONACYT-SENER) %< avec comité de lecture %@ 2161-5063 %J ACS Synthetic Biology %I American Chemical Society %V 7 %N 12 %P 2742-2749 %8 2018 %D 2018 %R 10.1021/acssynbio.8b00313 %M 30475588 %K NADPH regeneration %K auxotroph strain %K selection strain %K enzyme screening %K dihydrolipoamide dehydrogenase %K glyceraldehyde 3-phosphate dehydrogenase %K formate dehydrogenase %K cinnamyl alcohol %Z Life Sciences [q-bio]Journal articles %X Insufficient rate of NADPH regeneration often limits the activity of biosynthetic pathways. Expression of NADPH-regenerating enzymes is commonly used to address this problem and increase cofactor availability. Here, we construct an Escherichia coli NADPH-auxotroph strain, which is deleted in all reactions that produce NADPH with the exception of 6-phosphogluconate dehydrogenase. This strain grows on a minimal medium only if gluconate is added as NADPH source. When gluconate is omitted, the strain serves as a "biosensor" for the capability of enzymes to regenerate NADPH in vivo. We show that the NADPH-auxotroph strain can be used to quantitatively assess different NADPH-regenerating enzymes and provide essential information on expression levels and concentrations of reduced substrates required to support optimal NADPH production rate. The NADPH-auxotroph strain thus serves as an effective metabolic platform for evaluating NADPH regeneration within the cellular context. %G English %L cea-02291351 %U https://cea.hal.science/cea-02291351 %~ CEA %~ CNRS %~ UNIV-EVRY %~ DSV-IG %~ DSV %~ CEA-UPSAY %~ GENOMIQUE-METABOLIQUE %~ UNIV-PARIS-SACLAY %~ UNIV-EVRY-SACLAY %~ CEA-UPSAY-SACLAY %~ JACOB %~ CEA-DRF %~ GENOSCOPE %~ TEST-HALCNRS %~ GS-BIOSPHERA