期刊
ELIFE
卷 7, 期 -, 页码 -出版社
ELIFE SCIENCES PUBLICATIONS LTD
DOI: 10.7554/eLife.32638
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资金
- Max-Planck-Gesellschaft
- Ecole Polytechnique Federale de Lausanne
We introduce 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, we observe how cells can react to such changes by adapting metabolic fluxes. Finally, we demonstrate how a comparison of the effect of drugs on cellular NAD(P) levels can be used to probe mechanisms of action.
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