Location of Diphenylhexatriene (DPH) and Its Derivatives within Membranes: Comparison of Different Fluorescence Quenching Analyses of Membrane Depth

Abstract

The average membrane location of a series of diphenylhexatriene (DPH)-derived membrane probes was analyzed by measuring the quenching of DPH fluorescence with a series of nitroxide-labeled lipids in which the depth of the nitroxide group is varied. All DPH derivatives were located deeply within the bilayer. Some derivatives were anchored at a shallower depth than free DPH by attachment to cationic or anionic groups. However, the absolute change in DPH depth upon attachment to such groups was relatively modest (Methods Enzymol. 278, 462−473). Again more physically reasonable results were obtained from parallax analysis, likely due to non-Gaussian behavior of the depth dependence of quenching. Notwithstanding these observations, the significant number of cases where Gaussian curve fitting methods for quenching analysis are most powerful are discussed.