Evidence for the Extended Phospholipid Conformation in Membrane Fusion and Hemifusion

Abstract

Molecular-level mechanisms of fusion and hemifusion of large unilamellar dioleoyl phosphatidic acid/phosphocholine (DOPA/DOPC, 1:1 molar ratio) vesicles induced by millimolar Ca 2+ and Mg 2+, respectively, were investigated using fluorescence spectroscopy. In keeping with reduction of membrane free volume V f, both divalent cations increased the emission polarization for 1,6-diphenyl-1,3,5-hexatriene (DPH). An important finding was a decrease in excimer/monomer emission intensity ratio ( I e/ I m) for the intramolecular excimer-forming probe 1,2-bis[(pyren-1-)yl]decanoyl-sn-glycero-3-phosphocholine (bis-PDPC) in the course of fusion and hemifusion. Comparison with another intramolecular excimer-forming probe, namely, 1-[(pyren-1)-yl]decanoyl-2-[(pyren-1)-yl]tetradecanoyl-sn-glycero-3-phosphocholine (PDPTPC), allowed us to exclude changes in acyl chain alignment to be causing the decrement in I e/ I m. As a decrease in V f should increase I e/ I m for bis-PDPC and because contact site between adhering liposomes was required we conclude the most feasible explanation to be the adoption of the extended conformation (P.K.J. Kinnunen, 1992, Chem. Phys. Lipids 63:251–258) by bis-PDPC. In this conformation the two acyl chains are splaying so as to become embedded in the opposing leaflets of the two adhered bilayers, with the headgroup remaining between the adjacent surfaces. Our data provide evidence for a novel mechanism of fusion of the lipid bilayers.