Biomedical subjects

R Gorges

Publications and source records attributed to R Gorges.

5 recordsLinked to original sources

PubMed2000

Lateral microheterogeneity of diphenylhexatriene-labeled choline phospholipids in the erythrocyte ghost membrane as determined by time-resolved fluorescence spectroscopy.

Choline phospholipids are the major constituents of the outer layer of the erythrocyte membrane. To investigate their lateral membrane organization we determined the fluorescence lifetime properties of diphenylhexatriene analogues of phosphatidylcholine, choline plasmalogen, (the respective enolether derivative), and sphingomyelin inserted into the outer layer of hemoglobin-free ghosts. Fluorescence lifetimes were recorded by time-resolved phase and modulation fluorometry and analyzed in terms of Continuous Lorentzian distributions. To assess the influence of membrane proteins on the fluorescence lifetime of the labeled lipids in the biomembrane, lipid vesicles were used as controls. In general, the lifetime distributions in the ghost membranes are broad compared to vesicles. Phosphatidylcholine and sphingomyelin exhibit very similar lifetime distributions in contrast to an increased plasmalogen lifetime heterogeneity in both systems. Orientational effects of side chain mobilities on the observed lifetimes can be excluded. Fluorescence anisotropies revealed identical values for all three labeled phospholipids in the biomembrane.

Choline↗

PubMed1995

Transfer of phospholipase A-resistant pyrene-dialkyl-glycerophosphocholine to plasma lipoproteins: differences between Lp[a] and LDL.

1-O-Hexadecyl-2-O-pyrenedecanyl-sn-glycero-3-phosphocholine, a non-hydrolyzable fluorescent diether analog of phosphatidylcholine (PC), was synthesized as a probe for studying phospholipid transfer to different lipoprotein classes with potential phospholipase activities. After incubation of total human plasma with the new probe at 37 degrees C for 4.5 h, a characteristic partition between the main lipoprotein fractions was observed. The fluorescent lipid was not degraded under these conditions and, therefore, served as a measure for choline glycerophospholipid distribution between plasma lipoproteins. In low density lipoprotein (LDL) and high density lipoprotein-3 (HDL3) the fluorescent PC analog showed only monomer fluorescence, whereas in Lp[a] and HDL2 monomer and excimer fluorescence were observed, indicating that the fluorescent phosphatidylcholine analog was incorporated into the respective lipoproteins to a different extent. According to the increased pyrene excimer fluorescence in Lp[a] compared with LDL the labeled phosphatidylcholine must be enriched and/or clustered in Lp[a]. Data from phospholipid and total fluorescence analyses are compatible with the assumption of higher label concentration in Lp[a]. On the other hand, transfer rates for serum protein-catalyzed lipid transport into isolated Lp[a] were slower as compared to LDL. It is suggested that slower lipid transfer to Lp[a] under these conditions is due to the decreased lipid mobility in the Lp[a] surface, whereas the higher extent of label partition into Lp[a] as observed in total plasma might be due to the higher affinity of apolipoproteins for phosphatidylcholine in Lp[a] (Sommer, A., et al. 1992. J. Biol. Chem. 267: 24217-24222). The use of a fluorescent dialkyl- instead of diacyl-glycerophosphocholine for transfer studies was mandatory, as we found that lipoproteins contained phospholipase A2 activity toward long-chain phosphatidylcholine. The lipoprotein-associated phospholipase A2 was three times more active in Lp[a] than in LDL. The degradation products formed by the phospholipase, fatty acids, and lyso-PC may add to the high atherogenic potential of Lp[a].

Biological Transport↗

PubMed1994

Apoprotein-phospholipid interactions in Lp(a).

Lipoprotein (a) (Lp(a)) and low-density lipoprotein (LDL) are structurally related to each other. Both exhibit identical phospholipid compositions and possess one molecule of apoprotein B-100 (apoB). Lp(a) contains, in addition, apoprotein (a) (apo(a)), which localizes to the particle surface and interacts with the apoB component by non-covalent and covalent forces. Protein-protein interaction is probably interrelated with protein-lipid interaction. Fluorescent analogs of phosphatidylcholine and sphingomyelin were inserted into the surface layer of LDL and Lp(a). The obtained fluorescence data reflecting mobility and distributional heterogeneity of the labeled lipids provided evidence that apo-proteins discriminate between choline phospholipids and preferentially associate with phosphatidylcholine. This effect is enhanced in Lp(a) because of the presence of apolipoprotein (a). Higher affinity for Lp(a) as compared with LDL was also observed with a fluorescent diether analog of phosphatidylcholine in native serum. In contrast, the time-dependent transfer of the same lipid into Lp(a) was slower compared with LDL, probably as a consequence of the more rigid surface of the former lipoprotein.

Apolipoproteins↗

PubMed1992

Organization of phosphatidylcholine and sphingomyelin in the surface monolayer of low density lipoprotein and lipoprotein(a) as determined by time-resolved fluorometry.

Fluorescent analogs of phosphatidylcholine (PC) and sphingomyelin (SM) labeled with diphenylhexatrienylpropionic acid (DPH) were prepared and incorporated into the surface layer of human low density lipoprotein (LDL) and lipoprotein(a) (Lp(a)). Fluorescence anisotropy measurements of DPH-PC and DPH-SM in both lipoprotein classes were carried out at different temperatures ranging from 20 to 37 degrees C. DPH-PC as well as DPH-SM were shown to reside in more rigid domains in Lp(a) than in LDL according to higher anisotropy values in Lp(a). In both LDL and Lp(a), DPH-PC experienced a more rigid environment than DPH-SM, suggesting different environments of PC and SM in the surface shell of the lipoproteins. Fluorescence lifetimes of the labeled lipoproteins were determined by phase and modulation fluorometry. We found bimodal Lorentzian distributions for the decay times of DPH-PC and DPH-SM in LDL and Lp(a). Lifetime distribution centers for labeled lipids were very similar except for DPH-PC in Lp(a) which was shifted to longer lifetimes, suggesting a less polar environment of PC in Lp(a) than in LDL. The distributional width of DPH-PC in Lp(a) was broader than in LDL. Accordingly, phosphatidylcholine must be localized in a more homogeneous environment in LDL as compared with Lp(a). On the other hand, no difference in distributional widths was observed for DPH-SM in both lipoproteins, showing that SM organization in Lp(a) is unaffected by apo(a). From the obtained fluorescence data we propose that apoproteins discriminate between the choline phospholipids and preferentially associate with phosphatidylcholine. This effect is enhanced in Lp(a) due to the presence of apolipoprotein(a).

Fluorescence Polarization↗

PubMed1991

Sulfhydryl-selective fluorescence labeling of lipoprotein(a) reveals evidence for one single disulfide linkage between apoproteins(a) and B-100.

Human lipoprotein(a) and low-density lipoprotein were labeled with two different sulfhydryl-selective fluorescence markers. The hydrophilic fluorophore lucifer yellow iodoacetamide and the apolar compound 6-acryloyl-2-(dimethylamino)naphthalene were used to derivatize free -SH groups in the lipoproteins. Three sulfhydryls could be detected in low-density lipoprotein, whereas only two cysteines were available in lipoprotein(a). One of the three -SH groups in low-density lipoprotein was shown to be located in close proximity to the particle surface. We suggest that this surface-exposed cysteine of apoprotein B-100 serves as a component for the disulfide linkage to apoprotein(a) in lipoprotein(a).

2-Naphthylamine↗