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Valuable information on protein-membrane organization may in principle be obtained from polarized-light absorption (linear dichroism, LD) measurement on shear-aligned lipid vesicle bilayers as model membranes. However, attempts to probe LD in the UV wavelength region (<250 nm) have so far failed because of strong polarized light scattering from the vesicles. Using sucrose to match the refractive index and suppress the light scattering of phosphatidylcholine vesicles, we have been able to detect LD bands also in the peptide-absorbing region (200 -230 nm). The potential of refractive index matching in vesicle LD as a general method for studying membrane protein structure was investigated for the membrane pore-forming oligopeptide gramicidin incorporated into the liposome membranes. In the presence of sucrose, the LD signals arising from oriented tryptophan side chains as well as from n3 * and 3 * transitions of the amide chromophore of the polypeptide backbone could be studied. The observation of a strongly negative LD for the first exciton transition (Ϸ204 nm) is consistent with a membrane-spanning orientation of two intertwined parallel gramicidin helices, as predicted by coupledoscillator theory.

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A series of ruthenium(II) complexes comprising differently substituted dipyridophenazine (dppz) ligands have been examined with respect to their orientation when bound to phosphatidylcholine liposome bilayers. The liposomes were subjected to shear flow, thereby deformed into ellipsoid shapes, and the orientation of chromophores in the bilayers could then be examined with polarized light (linear dichroism, LD). After correction for turbidity dichroism, LD distinguishes between chromophore transition moments oriented along the lipid chains (negative LD), and parallel to the membrane surface (positive LD). The polarizations, energies, and absorption overlap of the electronic transitions of the ruthenium complexes, known from previous work, could be used to characterize the orientation of liposome bound complexes. From luminescence lifetime studies further information about the localization of the chromophores was obtained. Assuming that the positively charged metal moiety of the complexes remains close to the bilayer surface, monomers with unsubstituted dppz as well as alkyl substituted dppz ligands were found to dip the dppz ligand down into the bilayer, whereas nitrile and amide substituents contributed to the alignment of the dppz group parallel to the surface. The findings give insights into mechanisms that govern orientation of membrane solutes, thus providing useful leads, for example, for the composition of membrane molecular devices.

The behavior of poly-γ-methyl-l-glutamate (pMeE) at the air−water interface has been studied with the surface film balance technique. In addition, Langmuir−Blodgett (LB) films of pMeE deposited on mica and quartz have been studied by atomic force microscopy (AFM) and circular and linear dichroism (CD and LD) spectroscopy. Depending on the spreading solvent, pMeE displays strikingly different compression isotherms. When spread from chloroform or trifluoroacetic acid (TFA) the surface pressure isotherms are consistent with that of a peptide in α-helix conformation. However, the latter solvent gives rise to isotherms with a considerably smaller apparent mean molecular area, A 0. When spread from pyridine, on the other hand, pMeE yields an isotherm that is expanded and inconsistent with the presence of a monolayer consisting entirely of α-helical peptides. Isotherms and AFM images strongly suggest that peptide aggregation and solvent retention are the main factors behind the isotherm differences. When the water-soluble spreading solvent TFA is used, pMeE forms discrete wormlike aggregates embedded in a monolayer matrix. In the pyridine case, aggregation in the spreading solvent and retention of pyridine in the film result in a rough aggregate network coexisting with discrete aggregates. No aggregation takes place when chloroform is used as spreading solvent. CD and LD spectra of the LB films reveal a pronounced lateral orientation of the α-helices in films spread from chloroform and TFA, while spectra of films spread from pyridine are consistent with unordered peptide strands in β-sheet conformation. In conclusion, the results show that if water-soluble and/or low-volatile solvents are used as spreading media, hydrophobic peptides cannot, a priori, be assumed to form proper monolayers.

Background : Human Rad51 protein (HsRad51) is a homologue of Escherichia coli RecA protein, and involved in homologous recombination. These eukaryotic and bacterial proteins catalyse strand exchange between two homologous DNA molecules, each forming a complex with single-stranded DNA (ssDNA) and ATP as the initial step. Both proteins hydrolyse ATP; however, the role of ATP hydrolysis appears to vary between the two proteins.

Efficient cellular uptake is crucial for the success of any drug directed towards targets inside cells. Peptide nucleic acid (PNA), a DNA analog with a promising potential as a gene-directed drug, has been shown to display slow membrane penetration in cell cultures. We here used liposomes as an in vitro model of cell membranes to investigate the effect on penetration of a PNA molecule colvalently modified with a lipophilic group, an adamantyl moiety. The adamantyl attachment was found to increase the membrane-penetration rate of PNA three-fold, as compared to corresponding unmodified PNA. From the penetration behaviour of a number of small and large molecules we could conclude that passive diffusion is the mechanism for liposome-membrane passage. Flow linear dichroism (LD) of the modified PNA in presence of rod-shaped micelles, together with octanol-water distribution experiments, showed that the adamantyl-modified PNA is amphiphilic; the driving force behind the observed increased membrane-penetration rate appears to be an accumulation of the PNA in the lipid double layer.

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