Two-dimensional (2-D) crystals of annexin V, grown by specific binding to phosphatidylserine-containing planar lipid films, were studied by electron image analysis. Images of negatively stained two-dimensional crystals showed diffraction peaks extending to 11 A. After correcting lattice distorsions and averaging over several crystalline areas, the resolution of the analysis was extended up to 8 A. Observed along a direction perpendicular to the membrane plane, the four homologous domains characteristic of annexin V exhibit a noticeable difference in their distribution of protein density. An unambiguous assignment of the domains was possible due to the similar packing of annexin V molecules in the 2-D crystals and in a 3-D crystal form with pseudo-R3 symmetry. The domains I and IV (numbering according to Huber et al., Embo J., 1990, 9, 3867-3874) appear well resolved. On the other hand, the two other domains, II and III, present an almost continuous density, with a protrusion extending outwards the annexin V molecule. In addition, no hydrophilic opening is resolved at the center of the molecule, yet a stain-filled 13-A structure is present, surrounded by domains I, II, and IV and distant by 5 A from the center of the molecule. We interpret these structural features as reflecting a conformational change in the annexin V structure resulting from its membrane binding.