Biochemical and morphological properties of membranes of unsaturated fatty acid auxotrophs of Salmonella typhimurium: effects of fluorinated myristic acids.

In order to investigate the utility of the fluorine-19 nucleus as a spectroscopic probe, a fluorinated analog of myristic acid has been incorporated into the membrane lipids of an unsaturated fatty acid auxotroph of Salmonella typhimurium. It is capable of supporting limited growth at temperatures above 37 degrees C. Freeze-fracture electron microscopic examinations of the membrane ultrastructure show a temperature and fatty acid supplement-dependent segregation of intramembranous protein particles into distinct patches in the auxotrophic membrane leaving intramembranous protein-denuded areas. The occurrence of these patches seems to be related to the phase separation of membrane lipids. Corresponding changes in the transport and accumulation of methyl thio-beta-D-galactopyranoside and tetracycline are observed. However, transport of histidine does not appear to be dependent on the physical state of the membrane lipids. The auxotroph shows differences in growth and morphological characteristics from those of the wild type. Functions of both inner and outer membranes are shown to be affected as a response to the fatty acid chain composition of the lipids.

Electron microscopic observation of domain movement in reconstituted erythrocyte membranes.

The movement of labeled concanavalin A (Con A) receptors in reconstituted human erythrocyte membranes was observed directly in an electron microscope, using an environmental stage that kept the sample fully hydrated at all experimental temperatures. Human erythrocyte membrane ghosts were spread on the air/water interface in a Langmuir trough. The surface monolayer film contained most native proteins and lipids of the erythrocyte membrane. The Con A receptors in the film were labeled with Con A-conjugated, 25-nm-diameter gold microspheres. Unsupported bilayer membranes were reconstituted by dipping a 1000-mesh grid through the labeled surface film. The reconstituted membrane samples were observed under low beam current and photographed by timed exposures with sensitive x-ray films. The total radiation per exposure was kept below the damage threshold of 5 X 10(-4) coulomb/cm2. The Con A-gold labels were observed to move in unison within local areas (domains) of the reconstituted membrane. The size of the domains and the velocity of the labels were measured as functions of temperature. The typical domain size was 10 micron2 and the typical velocity of the labels was 7 nm/sec. The minimum domain size and velocity were found at 17 degrees-28 degrees C. Reduction of the amount of cholesterol in the precursor erythrocyte membrane caused the domain velocity at 7 degrees C to decrease and the domain size to increase; the opposite effect was observed with cholesterol enrichment. The results indicate that the components of the erythrocyte membrane tended to form moving domains and that the motion was related to lipid phase separation in the bilayer.

Formation of water-soluble complex between the 1-34 fragment of parathyroid hormone and dimyristoylphosphatidylcholine.

Two biologically active, 34 amino acid fragments of parathyroid hormone interact with dimyristoylphosphatidylcholine to form lipoprotein particles. In the lipid-bound form these parathyroid hormone peptides exhibit an increased amount of folded secondary structure and the tryptophan residue of [Nle8, Nle18, Tyr34] b PTH (1-34) amide appears to become buried in a more hydrophobic environment. The lipoprotein particle which is formed has dimensions of approximately 65 X 7 nm but aggregates to larger structures with increasing temperature. Above the phase transition of the phospholipid the peptides no longer affect the morphology of the lipid and the spectral properties of the peptide are not perturbed by the lipid. This is similar to the behavior of glucagon with dimyristoylphatidylcholine. The results indicate that several nonhomologous peptide hormones have common features which allow them to fold into an amphipathic helix and solubilize phospholipid.

Lateral interaction of cholesterol in diacylphosphatidylcholesterol bilayers.

Thermotropic phase-transition properties of the aqueous dispersions of several diacylphosphatidylcholesterol (DRCh) analogs are examined. The aqueous dispersions of their calcium salts exhibit characteristic endothermic thermotropic transitions due to a change in the conformation of acyl chains. These dispersions consist of osmotically intact liposomes that trap ions, and at the transition temperature there is anomalous increase in the ion leakage. Wide-angle electron diffraction studies of DPCh . Ca monolayers also exhibit a transition from a sharp 4.25 A band to a broad one centering at 4.7 A, reflecting an order-disorder transition in the acyl chains. The long-range order in the organization of acyl chains of DRCh molecules could arise from intermolecular interactions between the cholesterol moieties to form a functional dimer, and such dimers are apparently cross-linked by Ca2+ to form a long-range interacting lattice of acyl chains. Evidence for this model is adduced from the fluorescence properties of the dispersions of dimyristoylphosphatidylcholesta-5,7,9-trienol. The phase-transition properties of DRCh are an ideal illustration of calcium-induced isothermal phase transition.