  Annexins are a family of proteins that bind to anionic phospholipid membranes
in a Ca2+-dependent manner. Annexin A2 forms heterotetramers (Anx A2t) with the
S100A10 (p11) protein dimer. The tetramer is capable of bridging phospholipid
membranes and it has been suggested to play a role in Ca2+-dependent exocytosis
and cell-cell adhesion of metastatic cells. Here, we employ x-ray reflectivity
measurements to resolve the conformation of Anx A2t upon Ca2+-dependent binding
to single supported lipid bilayers (SLBs) composed of different mixtures of
anionic (POPS) and neutral (POPC) phospholipids. Based on our results we
propose that Anx A2t binds in a side-by-side configuration, i.e., both Anx A2
monomers bind to the bilayer with the p11 dimer positioned on top. Furthermore,
we observe a strong decrease of lipid mobility upon binding of Anx A2t to SLBs
with varying POPS content. X-ray reflectivity measurements indicate that
binding of Anx A2t also increases the density of the SLB. Interestingly, in the
protein-facing leaflet of the SLB the lipid density is higher than in the
substrate-facing leaflet. This asymmetric densification of the lipid bilayer by
Anx A2t and Ca2+ might have important implications for the biochemical
mechanism of Anx A2t-induced endo- and exocytosis.
