New paper published in eLife describes a new class of semisynthetic fluorescent biosensors for measuring free (NAD+) and NADPH/ NADP+ in live cells. The sensors represent powerful tools to study the role of NAD(P) in metabolism and signalling in healthly and diseased cells. This publication stemming from Kai Johnsson’s work at the NCCR Chemical Biology describes the development of a sensor to measure NAD(P) in living cells based on the Snifit design principle.
The authors introduced a new class of semisynthetic fluorescent biosensors for the quantification of free nicotinamide adenine dinucleotide (NAD+) and ratios of reduced to oxidized nicotinamide adenine dinucleotide phosphate (NADPH/NADP+) in live cells. Sensing is based on controlling the spatial proximity of two synthetic fluorophores by binding of NAD(P) to the protein component of the sensor. The sensors possess a large dynamic range, can be excited at long wavelengths, are pH-insensitive, have tunable response range and can be localized in different organelles. Ratios of free NADPH/NADP+ are found to be higher in mitochondria compared to those found in the nucleus and the cytosol. By recording free NADPH/NADP+ ratios in response to changes in environmental conditions, the authors observed how cells can react to such changes by adapting metabolic fluxes. Finally, they demonstrated how a comparison of the effect of drugs on cellular NAD(P) levels can be used to probe mechanisms of action.