Unique Properties of Tightly Docked Membranes Prior to their Fusion

Abstract

Membrane merger through fusion is essential in myriad of processes including neurotransmission and viral infection. Although there are various classes of fusion proteins, they are believed to catalyze membrane fusion through a common pathway. Despite the importance of understanding of this process, the exact mechanisms underlying steps immediately before the first bilayer disruption remain uncharacterized. Here, we characterized mechanism leading to a change in lipid arrangement in membranes arrested in a state that precedes hemifusion. Such tightly adhering membranes with a distance of <1 nm are known from cryo-electron microscopy as fusion intermediates of trafficking organelles, mitochondria, or during cell entry by enveloped viruses. With the use of an in vitro reconstitution system consisting of lipid vesicles with SNARE proteins, we demonstrate formation of a metastable, divalent cation-independent, and protein-free membrane-membrane interface characterized by increased thickness. Furthermore, we confirm that structural rearrangements in the membranes are arising due to membrane intrinsic properties by performing all-atom molecular dynamics simulations of double-membrane systems at varying distances. In turn, we would like to propose that increase in membrane thickness arises by dehydration-driven lipid headgroup tilting leading to lateral area shrinkage that results in increased bilayer thickness visible in experiments.