Fluorescence quenching in model membranes: phospholipid acyl chain distributions around small fluorophores

Fluorescence quenching in lipid bilayers is treated by a new approach based on calculation of the probability distribution of quenching and nonquenching acyl chains around a fluorophore. The effect of acyl lattice site dependence (i.e., correlations of phospholipid sister chain occupancy of neighbor sites) was modeled by use of Monte Carlo simulations of acyl chain occupancy. This explicit accounting of site occupancy correlation was found to fit observed quenching behavior better than did a model wherein phospholipid quenchers are considered to be independent. A key aspect of this approach is to evaluate the rate for quenching in a bilayer composed of pure quenching lipid. In order to evaluate this quenching rate, and also to provide a strong test of the calculated probability distributions, we synthesized lipids with both acyl chains labeled with a quenching moiety (Br or nitroxide), as well as the more usual single-chain quenchers. The fluorescence of tryptophan octyl ester (TOE), and of the 1,6-diphenyl-1,3,5-hexatriene (DPH) derivatives trimethylammonium-DPH (TMA-DPH) and 1-lauroyl-2-(DPH-propionyl)phosphatidylcholine (DPH-PC), was examined. We obtained consistent results with all the fluorophores and quenchers indicating that up to 18 neighboring acyl sites can contribute to quenching, corresponding to two shells of acyl sites on a hexagonal lattice. Calculated discrete distributions of fluorescence intensities were converted into fluorescence lifetimes and compared with Gaussian and Lorentzian continuous lifetime distributions. The fluorescence quenching theory presented here may be used to explain quantitatively the heterogeneity of fluorophore environments in multicomponent membranes.