Interaction of a highly potent dimeric enkephalin analog, biphalin, with model membranes

Abstract

Biphalin, (Tyr-D-Ala-Gly-Phe-NH)2, is a highly potent dimeric analog of enkephalin. Its analgesic efficacy is due in part to its ability to permeate the blood-brain barrier. To aid in understanding the mechanism of the transmembrane movement we determined and analyzed the permeability and partition coefficients of biphalin and a series of analogues where F, Cl, I, NO2, or NH2 were placed in the para position of the aromatic rings of Phe4,4'. Liposomes composed of neutral phospholipids and cholesterol were used as the model membrane. The overall good correlation between permeability and water-membrane partition coefficients suggests that the movement of biphalins across the model membrane is controlled by diffusion and depends on the water-membrane partition coefficient. To explain the observed correlation between permeability and the electron withdrawing/donating character of the substituents in the phenylalanine ring, we examined various folding patterns of Leu-enkephalin, an endogenous pentapeptide that exhibits affinities toward the same classes of opioid receptors (delta and mu). The observed permeabilities and partition coefficients of biphalin and analogues, as well as the tyrosine side chain accessibility, are consistent with the presence of the type of folding where the tyrosine and phenylalanine side chains are in a close contact. We propose that the aromatic ring interaction can promote the peptide permeability by stabilizing a more compact structure of biphalin that would minimize the number of hydrogen bonds with water and therefore enhances partitioning into the model membrane.

The non-opioid receptor, antioxidant properties of morphine and the opioid peptide analog biphalin

“…However, to act intracellularly, biphalin must penetrate the cellular membrane. Biphalin may partially enter neuronal cells via simple diffusion [7]. However, opioid receptors may selectively transport this opioid ligand through the cell membrane.…”

“…The discovery of the opioid receptor system led to the belief that all of the effects of opioids were associated with their receptors' activation [2,8]. However, opioid drugs, such as morphine or fentanyl, or endogenous opioid peptides, such as enkephalins or endomorphins, may non-specifically interact with a number of macromolecules and biological membranes [7]. Abundant non-specific binding of opioid ligands has been observed even in simple in vitro opioid receptor binding experiments [non published observation].…”

Biomimetic Model Membrane Systems Serve as Increasingly Valuable in Vitro Tools

“…For the permeability assay, the sample of liposome encapsulated peptides were dialyzed and the amount of peptide released into the buffer was determined by a fluorescence assay, which measured the reaction product from the peptide amino group with fluorescamine (ex380/em475) (Romanowski et al, 2002). Partition coefficients were obtained using equilibrium dialysis experiments and the fluorescence measurements of the peptide Tyr and Trp residues (Romanowski et al, 1997). Interestingly, increased surface charge by addition of POPS showed increased biphalin binding but reduced translocation of the peptide across the bilayer.…”

Permeability of Phospholipid Vesicles to the Tumor Antigen Epitope gp100280–288

“…As far as the lipid is concerned, a number of parameters [27] affects vesicle permeability to peptides: notably, increasing the aliphatic chain length slightly reduces permeability; [28] the presence of cholesterol decreases membrane permeability [18] (but see reference [29]), whereas the presence of polymer-conjugated lipids should increase this parameter, [19] although opposite results have also been reported. [30] In addition, a number of other parameters, such as lipid concentration, sonication time, and even the shape of the preparation vial [23] affects vesicle size, size distribution, and the number of lipid layers, modifying permeability.…”

“…[18,27] Thus, it appears wise to assume that vesicle permeability to class I-restricted epitopes is peptide-and lipid-specific, and that both the lipid and the peptide side must be taken into account. However, it is also possible to speculate that modifications of these factors may have a limited impact on the use of peptidic epitopes as vaccines.…”