Spontaneous Transfer and Molecular Organization of Dehydroergosterol in Lipid Bilayers
Dehydroergosterol (DHE), a fluorescent cholesterol analogue, was used as a cholesterol mimic to study both the transfer kinetics between phospho-lipid bilayers and the molecular organization of this sterol in the bilayer vesicles during transfer. The fluorescence intensity was used to monitor the transfer rate of DHE between 1-palmitoyl-2-oleoyl-phosphatidylcholine small unilamellar vesicles at different temperatures. The equation that best fits the data is a single exponential decay plus a base value due to a non-exchangeable pool, A exp(-kt)+B. The fitted halftimes (t½ = ln2/k) and base values (B) are: 61 min and 45.4 at 16°C., 27 min and 25.1 at 24°C., 18 min and 20.0 at 37°C. These values are comparable to those obtained using 3H-labeled cholesterol (Bar et al., 1986), suggesting that DHE resembles cholesterol and that a non-exchangeable pool exists for DHE as well as cholesterol. In order to examine the changes in DHE organization during the transfer, acrylamide quenching was performed on the donor vesicles both at the beginning of transfer, designed as t°, and at the end of transfer, t∞. Our results show that the quenching rate constant at t° is greater than that at t∞. However, three days after the transfer, no apparent difference in the quenching rate constant was observed between t° and t∞. This indicates that a reorganization of DHE occurs within this time period. The fluorescence lifetime is 0.88 ns and 0.76 ns for t° and t∞, respectively. Rotational correlation times at t° and t∞ also exhibit a small difference.
KeywordsLipid Bilayer Fluorescence Lifetime Molecular Organization Rotational Correlation Time Quench Rate Constant
- Bar, L. K., Barenholz, Y., and Thompson, T. E., (1986) Biochemistry, 26:6701.Google Scholar