Peptide mimics of SNARE transmembrane segments drive membrane fusion depending on their conformational plasticity

SNARE proteins are essential for different types of intracellular membrane fusion. Whereas interaction between their cytoplasmic domains is held respo nsible for establishing membrane proximity, the role of the transmembrane s egments in the fusion process is currently not clear. Here, we used an in v itro approach based on lipid mixing and electron microscopy to examine a po tential fusogenic activity of the transmembrane segments. We show that the presence of synthetic peptides representing the transmembrane segments of t he presynaptic soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARES) synaptobrevin II (also referred to as VAMP II) or synta xin 1A, but not of an unrelated control peptide, in liposomal membranes dri ves their fusion. Liposome aggregation by millimolar Ca2+ concentrations st rongly potentiated the effect of the peptides; this indicates that juxtapos ition of the bilayers favours their fusion in the absence of the cytoplasmi c SNARE domains. Peptide-driven fusion is reminiscent of natural membrane f usion, since it was suppressed by lysolipid and involved both bilayer leafl ets. This suggests transient presence of a hemifusion intermediate followed by complete membrane merger. Structural studies of the peptides in lipid b ilayers performed by Fourier transform infrared spectroscopy indicated mixt ures of a-helical and P-sheet conformations. In isotropic solution, circula r dichroism spectroscopy showed the peptides to exist in a concentration-de pendent equilibrium of a-helical and P-sheet structures. Interestingly, the fusogenic activity decreased with increasing stability of the a-helical so lution structure for a panel of variant peptides. Thus, structural plastici ty of transmembrane segments may be important for SNARE protein function at a late step in membrane fusion. (C) 2001 Academic Press.