Semisynthetic biosensors for mapping cellular concentrations of nicotinamide adenine dinucleotides
Olivier Sallin,
Luc Reymond,
Corentin Gondrand,
Fabio Raith,
Birgit Koch,
Kai Johnsson
Affiliations
Olivier Sallin
École Polytechnique Fédérale de Lausanne, Institute of Chemical Sciences and Engineering, Lausanne, Switzerland
Luc Reymond
École Polytechnique Fédérale de Lausanne, Institute of Chemical Sciences and Engineering, Lausanne, Switzerland; National Centre of Competence in Research in Chemical Biology, Lausanne, Switzerland
Corentin Gondrand
Department of Chemical Biology, Max-Planck-Institute for Medical Research, Heidelberg, Germany
École Polytechnique Fédérale de Lausanne, Institute of Chemical Sciences and Engineering, Lausanne, Switzerland; Department of Chemical Biology, Max-Planck-Institute for Medical Research, Heidelberg, Germany; National Centre of Competence in Research in Chemical Biology, Lausanne, Switzerland
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.
