Novel Organization and Properties of Annexin 2-Membrane Complexes

Abstract Annexin 2 belongs to the annexin family of proteins that bind to phospholipid membranes in a Ca2+-dependent manner. Here we show that, under mild acidic conditions, annexin 2 binds to and aggregates membranes containing anionic phospholipids, a fact that questions the mechanism of its interaction with membranes via Ca2+ bridges only. The H+ sensitivity of annexin 2-mediated aggregation is modulated by lipid composition (i.e. cholesterol content). Cryo-electron microscopy of aggregated liposomes revealed that both the monomeric and the tetrameric forms of the protein form bridges between the liposomes at acidic pH. Monomeric annexin 2 induced two different organizations of the membrane junctions. The first resembled that obtained at pH 7 in the presence of Ca2+. For the tetramer, the arrangement was different. These bridges seemed more flexible than the Ca2+-mediated junctions allowing the invagination of membranes. Time-resolved fluorescence analysis at mild acidic pH and the measurement of Stokes radius revealed that the protein undergoes conformational changes similar to those induced by Ca2+. Labeling with the lipophilic probe 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)diazirine indicated that the protein has access to the hydrophobic part of the membrane at both acidic pH in the absence of Ca2+ and at neutral pH in the presence of Ca2+. Models for the membrane interactions of annexin 2 at neutral pH in the presence of Ca2+ and at acidic pH are discussed.