The physical properties of the core and the surface of five classes of human plasma
lipoproteins were investigated using five
fluorescent probes. The location of the fluorescence probes in the
lipoprotein assembly was determined using collisional quenching and resonance energy transfer. The fluorophores monitor different regions of the
lipoproteins, as shown by fluorescence quenching.
Diphenylhexatriene (DPH) and methyl trans-
parinaric acid (MTPA), which are apolar molecules, are localized mainly in the
lipoprotein core. Their distribution into the surface is dependent upon the volume ratio of the hydrophobic part of the envelope and the core. The polar fluorophores, trimethylaminodiphenylhexatriene (TMADPH), hydroxycoumarin (HC) and trans-
parinaric acid (TPA) are anchored in the
glycerol skeleton region of the surface monolayer with the fluorophore group of HC in the headgroup region of the
phospholipids. We determined the temperature-dependent steady-state fluorescence anisotropy (r) of these fluorophores in the four major classes of
lipoproteins: VLDL,
LDL, HDL2, HDL3 and in abnormal HDL from
abetalipoproteinemia patients (HDLab). The hydrophobic probes, DPH and MTPA, reported the r values in the
lipoproteins in the following order:
LDL greater than HDL2 greater than HDL3 much greater than VLDL. This order correlates with the
triglyceride-to-
cholesterol ester (TG/CE) ratio in the core of
lipoproteins. The polar probes HC, TPA and TMADPH reported the r value in a different order: HDL2, HDL3 greater than or equal to
LDL much greater than VLDL. This is compatible with the decreasing order of the
protein to
lipid ratio in the envelope of these
lipoproteins. HDLab was investigated by three
fluorescent probes: DPH, TMADPH and HC. The anisotropy of DPH in HDLab was larger than that of either HDL2 or HDL3 in normal donors, probably due to the smaller TG/CE ratio in HDLab. The lower r values reported by HC and TMADPH for HDLab are not fully understood and may be related to other factors such as acyl chains composition. The characterization of
lipoproteins by fluorescence depolarization using probes of known location in the
lipoprotein assembly is very sensitive and may be used to report deviation from the norm.