gramicidin-a and 1-2-dielaidoylphosphatidylethanolamine

Addition of short-chain phospholipids to the gramicidin S-DNA-dioleoyl phosphatidylethanolamine complex enhanced up to 6-fold beta-galactosidase expression in several cell-lines in vitro. Among the compounds tested, the most potent in enhancing transfection were the dicapryl- and the dicapryloyl phosphatidylcholine. In contrast, no significant enhancement of transfection was seen when short-chain phospholipids were mixed with cationic lipids. Short-chain phospholipid and gramicidin S may act simultaneously on the cell membrane to enhance gene transfer, yet resulting in no toxicity.

Topics: Animals; beta-Galactosidase; Cell Line; Cell Membrane; DNA; Gene Transfer Techniques; Gramicidin; Humans; Mice; Phosphatidylcholines; Phosphatidylethanolamines; Phospholipids; Structure-Activity Relationship

Gramicidin-lipid interactions were investigated using diacylphosphatidylcholines that contained two identical acyl chains of varying length, between 6 and 14 carbons. The gramicidin A (gA) conformation was monitored by circular dichroism (CD) spectroscopy and high-performance size-exclusion chromatography, and the lipid organization was investigated using 31P and 1H NMR spectroscopy and negative-stain electron microscopy. Diacylphosphatidylcholine (PC) lipids with chain lengths between 4 and 8 carbons have been previously shown to have a micellar organization in aqueous solution [Lin, T.-L., et al. (1986) J. Am. Chem. Soc. 108, 3499-3507]. CD spectra of aqueous gA/lipid dispersions, at a ratio of 1:28, demonstrated that the channel conformation of gA can be readily obtained when the acyl chain length is > or = 10, but not when the chain length is < or = 7. Size-exclusion chromatography revealed that the fraction of gA that could easily be dissociated into monomers in the dispersions increased with increasing acyl chain length, in agreement with the CD results. For a chain length of 8, the results were intermediate. The formation of the channel structure was found to depend on the "solvent-history", the temperature, the gA and lipid concentrations, the gA:lipid ratio, and consequently on the method of sample preparation. 1H and 31P NMR results suggest that codispersed gA increases the size of dioctanoyl-PC aggregates, but not of dihexanoyl-PC micelles. Negative-stain electron microscopy directly supports these findings. Dihexanoyl-PC (28 mM) was able to solubilize 1 mM gA in H2O, but the gA was not in the "channel" conformation.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Amino Acid Sequence; Chromatography, Gel; Chromatography, High Pressure Liquid; Circular Dichroism; Gramicidin; Hydrogen; Magnetic Resonance Spectroscopy; Micelles; Microscopy, Electron; Molecular Sequence Data; Phosphatidylcholines; Phosphatidylethanolamines; Phosphorus Isotopes; Protein Conformation; Solubility

Gramicidin films at the air/water interface are shown to exhibit a phase transition at 225 A2/molecule which might be caused by either cluster formation, reorientation of molecules, conformational changes or multilayer formation. It is further shown that coupling of a charged group on either NH2- or COOH-terminus or elongation of the peptide by two amino acids, only slightly affects the surface area characteristics whereas modification of the tryptophans or even replacement of a single tryptophan by phenylalanine leads to drastic alterations in the surface-area characteristics and a (partial) loss of the phase transition demonstrating that the tryptophans play an important role in the interfacial behavior of gramicidin. The lack of a solvent history effect on the interfacial behavior indicates a rapid conformational interconversion of the peptide at the air/water interface. Gramicidin in mixtures with dioleoylphosphatidylcholine and lysopalmitoylphosphatidylcholine shows a condensing effect whereas gramicidin shows ideal mixing with dioleoylphosphatidylethanolamine. The condensing effect most likely is related to the aggregational state of the peptides which is different in phosphatidylcholines and phosphatidylethanolamines.

Topics: Amino Acid Sequence; Gramicidin; Lipid Bilayers; Models, Biological; Molecular Sequence Data; Phosphatidylcholines; Phosphatidylethanolamines; Pressure; Structure-Activity Relationship; Surface Properties

The following results are reported in this paper: The interaction of gramicidin with [11,11-2H2]dioleoylphosphatidylcholine (DOPC) and [11,11-2H2]dioleoylphosphatidylethanolamine (DOPE) at different stages of hydration was studied by 2H- and 31P-nuclear magnetic resonance. In the L alpha phase in excess water the acyl chains of phosphatidylethanolamine (PE) are more ordered than phosphatidylcholine (PC) most likely as the result of the lower headgroup hydration of the former lipid. In excess water gramicidin incorporation above 5 mol % in DOPC causes a bilayer----hexagonal HII phase change. In the HII phase acyl chain order is virtually unaffected by gramicidin but the peptide restricts the fast chain motions. At low water content gramicidin cannot induce the HII phase but it markedly decreases chain order in the DOPC bilayer. Increasing water content results in separation between a gramicidin-poor and a gramicidin-rich L alpha phase with decreased order of the entire lipid molecule. Further increase in hydration reverts at low gramicidin contents the phase separation and at high gramicidin contents results in a direct change of the disordered lamellar to the hexagonal HII phase. Gramicidin also promotes HII phase formation in the PE system but interacts much less strongly with PE than with PC. The results support our hypothesis that gramicidin, by a combination of strong intermolecular attraction forces and its pronounced cone shape, both involving the four tryptophans at the COOH-terminus, has a strong tendency to organize, with the appropriate lipid, in intramembranous cylindrical structures such as is found in the HII phase.

Topics: Deuterium; Gramicidin; Magnetic Resonance Spectroscopy; Models, Biological; Molecular Conformation; Phosphatidylcholines; Phosphatidylethanolamines; Thermodynamics