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Abstract
The fluorescent dye BADAN (6-bromoacetyl-2-dimetylaminonaphtalene) is widely used in various fields of life sciences, however, the photophysical properties of BADAN are not fully understood. The study of the spectral properties of BADAN attached to a number of mutant forms of GGBP, as well as changes in its spectral characteristics during structural changes in proteins, allowed to shed light on the photophysical properties of BADAN. It was shown that spectral properties of BADAN are determined by at least one non-fluorescent and two fluorescent isomers with overlapping absorbing bands. It was found that BADAN fluorescence is determined by the unsolvated “PICT” (planar intramolecular charge transfer state) and solvated “TICT” (twisted intramolecular charge transfer state) excited states. While “TICT” state can be formed both as a result of the “PICT” state solvation and as a result of light absorption by the solvated ground state of the dye. BADAN fluorescence linked to GGBP/H152C apoform is quenched by Trp 183, but this effect is inhibited by glucose intercalation. New details of the changes in the spectral characteristics of BADAN during the unfolding of the protein apo and holoforms have been obtained.
Highlights
Published: 15 October 2021Thanks to high sensitivity, simplicity, low cost, and possibility to be used as an express test, fluorescence methods are widely used in the study and analysis of living systems.For different tasks, researchers have a wide range of fluorescent methods in their arsenal; namely, intrinsic fluorescence, fluorescent proteins, external fluorescent probes, dyes, and so on
We studied the spectral characteristics of BADAN linked to glucose/D-galactose-binding protein (GGBP) mutants (GGBP/H152C, GGBP/W284C, GGBP/H152C/W183F, and GGBP/H152C/W183A)
In the mutant form GGBP/H152C, BADAN is located in close proximity to the glucose binding site
Summary
Researchers have a wide range of fluorescent methods in their arsenal; namely, intrinsic fluorescence, fluorescent proteins, external fluorescent probes, dyes, and so on. PRODAN and LAURDAN are known to be used in several membrane studies. Their photophysical properties have been extensively studied and the existence of several states has been established, of PRODAN and its analogs, in both the ground and excited states [1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18]. It was shown that the fluorescence characteristics of PRODAN in
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