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|Title: ||Fluorescence of nitrobenzoxadiazole (NBD)-labeled lipids in model membranes is connected not to lipid mobility but to probe location|
|Authors: ||Amaro, Mariana|
Filipe, Hugo A. L.
Prates Ramalho, J. P.
Loura, Luís M. S.
|Issue Date: ||2016|
|Citation: ||Amaro, Mariana; Filipe, Hugo A. L.; Prates Ramalho, J. P.; Hof, Martin; Loura, Luís M. S.Fluorescence of nitrobenzoxadiazole (NBD)-labeled lipids in model membranes is connected not to lipid mobility but to probe location, Phys. Chem. Chem. Phys. 18, 10, 7042-7054, 2016.|
|Abstract: ||Nitrobenzoxadiazole (NBD)-labeled lipids are popular fluorescent membrane probes. However, the
understanding of important aspects of the photophysics of NBD remains incomplete, including the
observed shift in the emission spectrum of NBD-lipids to longer wavelengths following excitation at
the red edge of the absorption spectrum (red-edge excitation shift or REES). REES of NBD-lipids in
membrane environments has been previously interpreted as reflecting restricted mobility of solvent
surrounding the fluorophore. However, this requires a large change in the dipole moment (Dm) of NBD
upon excitation. Previous calculations of the value of Dm of NBD in the literature have been carried out
using outdated semi-empirical methods, leading to conflicting values. Using up-to-date density
functional theory methods, we recalculated the value of Dm and verified that it is rather small (B2 D).
Fluorescence measurements confirmed that the value of REES is B16 nm for 1,2-dioleoyl-sn-glycero-3-
phospho-L-serine-N-(NBD) (NBD-PS) in dioleoylphosphatidylcholine vesicles. However, the observed
shift is independent of both the temperature and the presence of cholesterol and is therefore insensitive
to the mobility and hydration of the membrane. Moreover, red-edge excitation leads to an increased
contribution of the decay component with a shorter lifetime, whereas time-resolved emission spectra of
NBD-PS displayed an atypical blue shift following excitation. This excludes restrictions to solvent
relaxation as the cause of the measured REES and TRES of NBD, pointing instead to the heterogeneous
transverse location of probes as the origin of these effects. The latter hypothesis was confirmed by
molecular dynamics simulations, from which the calculated heterogeneity of the hydration and location
of NBD correlated with the measured fluorescence lifetimes/REES. Globally, our combination of theoretical
and experiment-based techniques has led to a considerably improved understanding of the photophysics
of NBD and a reinterpretation of its REES in particular.|
|Other Identifiers: ||1463-9076|
|Appears in Collections:||CQE - Publicações - Artigos em Revistas Internacionais Com Arbitragem Científica|
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