Engineering of an NADPH/NADP+ Redox Sensor in Yeast | AIChE

Engineering of an NADPH/NADP+ Redox Sensor in Yeast

Authors 

Pihl, T. P. B., Technical University of Denmark
Pedersen, K. R., Technical University of Denmark

Biosensors have emerged as powerful tools for timely and precise in vivo diagnosis and selection.  In particular, biosensors that can couple intercellular cues with downstream signaling responses are currently attracting major attention within health science and biotechnology.  In this study, we report the engineering and application of a genetically encoded NADPH/NADP+ redox biosensor in the budding yeast Saccharomyces cerevisiae.  Using the biosensor, we are able to monitor the NADPH redox state and experimentally validate genetic mutants predicted to have changed NADPH availability.  We also show that the sensor can regulate the expression of an NADP+-oxidoreductase to suppress NADPH deficiency observed in a mutant with severely impaired NADPH regeneration. Finally, we couple the biosensor with expression of novel native selector genes, which can be used for selection of cells with higher NADPH/NADP+ ratios. Taken together, we show that hitchhiking and rational engineering of native signaling components can be applied for engineering of biosensors applicable to both monitoring and timely regulation of cell redox state. Such biosensors should be of interest to the metabolic engineering community for the facile identification of platform strains with high cofactor requirements.