Effect of phospholipid composition on an amphipathic peptide-mediated poreformation in bilayer vesicles

To better understand the influence of phospholipid acyl-chain composition o n the formation of pores by cytotoxic amphipathic helices in biological mem branes, the leakage of aqueous contents induced by the synthetic peptide GA LA (WEAALAEALAE ALAEHLAEALAEALEALAA) from large unilamellar phospholipid ve sicles of various compositions has been studied. Peptide-mediated leakage w as examined at pH 5.0 from vesicles made of phosphatidylcholine (PC) and ph osphatidylglycerol (PG) with the following acyl-chain compositions: 1-palmi toyl-2-oleoyl (PO), 1,2-dioleoyl (DO), 1,2-dielaidoyl (DE), and 1,2-dipetro selinoyl (DPe). A mathematical model predicts and simulates the final exten ts of GALA-mediated leakage of 1-aminonaphthalene-3,6,8-trisulfonic acid (A NTS) and p-xylene-bis-pyridinium bromide (DPX) from 1-palmitoyl-2-oleoyl-ph osphatidylcholine/-palmitoyl-2-oleoyl-phosphatidylglycerol (POPC/POPG) and 1,2-dielaidoyl-sn-glycero-3-phosphocholine/1,2-dielaidoyl-phosphatidylglyce rol (DEPC/DEPG) liposomes at pH 5.0 as a function of peptide concentration in the bilayer, by considering that GALA pores responsible for this leakage have a minimum size of 10 +/- 2 monomers and are formed by quasiirreversib le aggregation of the peptide. With the phospholipid acyl-chain composition s tested, GALA-induced ANTS/DPX leakage follows the rank order POPC/POPG ap proximate to DEPC/DEPG > DPePC/DPePG > DOPC/DOPG. Results from binding expe riments reveal that this reduced leakage from DOPC/DOPG vesicles cannot be explained by a reduced binding affinity of the peptide to these membranes. As shown by monitoring the leakage of a fluorescent dextran, an increase in the minimum pore size also does not explain the reduction in ANTS/DPX leak age. The data suggest that surface-associated GALA monomers or aggregates a re stabilized in bilayers composed of phospholipids containing a cis unsatu ration per acyl chain (DO and DPe), while transbilayer peptide insertion is reduced. GALA-induced ANTS/DPX leakage is also decreased when the vesicles contain phosphatidylethanolamine (PE). This lends further support to the s uggestion that factors stabilizing the surface slate of the peptide reduce its insertion and subsequent pore formation in the bilayer.