Blacker, Thomas S;
Mistry, Nimit;
Plotegher, Nicoletta;
Westbrook, Elizabeth R;
Sewell, Michael DE;
Carroll, John;
Szabadkai, Gyorgy;
... Duchen, Michael R; + view all
(2025)
Redox-dependent binding and conformational equilibria govern the fluorescence decay of NAD(P)H in living cells.
FEBS Letters
10.1002/1873-3468.70125.
(In press).
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Abstract
When probed in living systems using fluorescence lifetime imaging microscopy (FLIM), the emission from reduced nicotinamide adenine dinucleotide (NADH) and its phosphorylated form NADPH have shown promise as sensitive intrinsic reporters of metabolism. However, an incomplete understanding of the biochemical processes controlling their fluorescence decay makes it difficult to draw unambiguous conclusions. Here, we utilised time‐resolved fluorescence anisotropy imaging to identify multiple enzyme binding configurations associated with lifetimes both longer and shorter than unbound NAD(P)H. FLIM, combined with mathematical and computational modelling, revealed that the redox states of the NAD and NADP pools control the steady‐state equilibrium of binding configurations, which in turn determines the observed fluorescence decay. This knowledge will be foundational to developing the accurate interpretation of NAD(P)H FLIM.
| Type: | Article |
|---|---|
| Title: | Redox-dependent binding and conformational equilibria govern the fluorescence decay of NAD(P)H in living cells |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1002/1873-3468.70125 |
| Publisher version: | https://doi.org/10.1002/1873-3468.70125 |
| Language: | English |
| Additional information: | Copyright © 2025 The Author(s). FEBS Letters published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies. This is an open access article under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
| Keywords: | FLIM, fluorescence, fluorescence anisotropy, fluorescence lifetime, metabolism, microscopy, NAD(P)H, NADH, NADPH |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Structural and Molecular Biology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10211918 |
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