Synthesis and some properties of constrained short-chain phosphatidylcholine analogues: (+)- and (-)-(1,3/2)-1-O-(phosphocholine)2,3-O- dihexanoylcyclopentane-1,2,3-triol

Reported herein is the synthesis of (+)- and (-)-(1,3/2)-1-O-(phosphocholine)-2,3-O-dihexanoylcyclopentane-1,2, 3-triol. These are the enantiomers of a contrained analogue of dihexanoylphosphatidylcholine in which the glycerol backbone is replaced by all-trans cyclopentane-1,2,3-triol. Evidence is presented to demonstrate that the (-)-enantiomer is a substrate for phospholipase A2 (PLA2) (Crotalus atrox) while the (+)-enantiomer is not. This strict enantiomeric (and positional) specificity was exploited in conjunction with a novel application of DEAE-cellulose column chromatography, to achieve racemic resolution with an excellent yield. The constrained backbone geometry, and the experimentally accessible critical micellar concentration (CMC) of these analogues should render them useful probes for assessing the contribution of substrate conformation and flexibility to the catalytic efficiency of PLA2.     

Adhesion of phospholipid vesicles to Chinese hamster fibroblasts: Role of cell surface proteins

The adhesion of artificially generated lipid membrane vesicles to Chinese hamster V79 fibroblasts in suspension was used as a model system for studying membrane interactions. Below their gel-liquid crystalline phase transition temperature, vesicles comprised of dipalmitoyl lecithin (DPL) or dimyristoyl lecithin (DML) absorbed to the surfaces of EDTA- dissociated cells. These adherent vesicles could not be removed by repeated washings of the treated cells but could be released into the medium by treatment with trypsin. EM autoradiographic studies of cells treated with[(3)H]DML or [(3)H]DPL vesicles showed that most of the radioactive lipids were confined to the cell periphery. Scanning electron microscopy and fluorescence microscopy further confirmed the presence of adherent vesicles at the cell surface. Adhesion of DML or DPL vesicles to EDTA-dissociated cells modified the lactoperoxidase-catalyzed iodination pattern of the cell surface proteins; the inhibition of labeling of two proteins with an approximately 60,000- dalton mol wt was particularly evident. Incubation of cells wit h (3)H-lipid vesicles followed by sodium dodecyl sulfate (SDS)- polyacrylamide gel electrophoresis showed that some of the (3)H-lipid migrated preferentially with these approximately 60,000-mol wt proteins. Studies of the temperature dependence of vesicle uptake and subsequent release by trypsin showed that DML or DPL vesicle adhesion to EDTA- dissociated cells increased with decreasing temperatures. In contrast, cells trypsinized before incubation with vesicles showed practically no temperature dependence of vesicle uptake. These results suggest two pathways for adhesion of lipid vesicles to the cell surface-a temperature-sensitive one involving cell surface proteins, and a temperature-independent one. These findings are discussed in terms of current models for cell-cell interactions.     

Separation of transport and metabolic steps in the uptake of succinate byKluyveromyces fragilis

Characteristics of succinate uptake were determined in glucose-, glycerol-, and succinate-grown cells of the succinate-utilizingKluyveromyces fragilis. Glucose represses the uptake when present during growth and inhibits it when present during uptake. Absence of glucose or presence of succinate or glycerol derepresses the uptake. The transport and subsequent metabolic steps can be efficiently separated by inhibiting the latter with carboxin.     

Significance of the lag phase in K1 killer toxin action on sensitive yeast cells

The minimum period between the addition of killer toxin K1 to sensitive yeast cells and the appearance of first cells stained with bromocresol purple indicating membrane damage, is approximately 20 min. The length of this lag phase depends strongly on toxin concentration, extending sharply at toxin levels lower than 60 lethal units (LU) per cell (about one-tenth of the toxin concentration necessary for saturating all surface receptors). As the binding of the toxin to the cell is virtually complete within 1 min, the rest of the lag phase reflects processes different from actual binding,e.g. combination of several toxin molecules to form a membrane ion channel or pore.     

Osmotic pressure: Thermodynamic basis and units of measurement

The theoretical basis and practical implications of osmotic pressure measurement are briefly discussed. Stress is laid on interconversions of various tabulated and practically used units and on practical examples of osmotic pressure calculation and determination, with special reference to possible pitfalls in osmotic pressure assessment and adjustment. Part I of a series of overviews on osmotic pressure and osmosis.