Orientation and Dynamics of a Novel Fluorescent Cholesterol Analogue in Membranes of Varying Phase

Fluorescent analogues of cholesterol offer a powerful approach for monitoring cholesterol behavior in membranes because of their high sensitivity, suitable time resolution, and multiplicity of measurable parameters. In this work, we have monitored the orientation and dynamics of a novel fluorescent cholesterol probe, 6-dansylcholestanol (DChol), in membranes of different phase type utilizing sensitive fluorescence techniques including the red-edge excitation shift (REES) approach. Our results show that fluorescence emission maximum, anisotropy, and lifetime of DChol are dependent on the phase of the membrane. Interestingly, DChol exhibits significant red-edge excitation shift (REES) that appear to depend on the phase of the membrane. Analysis of membrane penetration depth by the parallax method shows that the dansyl group of DChol is localized at the inter-facial region of the membrane (similar to 15.6 angstrom from the center of the bilayer). This is in excellent agreement with the previously reported location of cholesterol in fluid-phase membranes. We propose that DChol could be a potentially useful cholesterol analogue in future studies of model and biological membranes.