Atomic force microscopy (AFM) had revealed two different forms of PLY prepores in planar bilayers, rising 110 or 80 Å above the membrane surface (van Pee et al., 2016). The low form has the same height as the pore complex, but the lipid bilayer is still intact. We propose that in this lower prepore, D1, D2 and D4 have moved to their positions in the pore assembly, while D3 has not yet refolded and the long, membrane-spanning β-hairpins have not yet formed. We assign the tall form to an early prepore state, which then rearranges into the lower, late prepore state. The cross-section profile (Figure 6B,D) shows that the early prepore is essentially a ring-shaped side-by-side arrangement of soluble PLY monomers (Figure 6—figure supplement 1), similar to that in the crystal lattice (Lawrence et al., 2015; Marshall et al., 2015; van Pee et al., 2016). While the late prepore was observed for 13% of the rings on planar bilayers examined by AFM, it was not observed in tomographic volumes of PLY on liposomes, implying that it is an intermediate, transient state which inserts more readily into curved liposomes than into planar bilayers. Cross-sections of pore complexes in liposomes (Figure 6B,E) show that the lipid bilayer around the prepores and pores is curved, whereas in the AFM images it was flat (van Pee et al., 2016). Prepore stability and membrane insertion thus seem to be related to membrane curvature, such that pores form preferentially in lipid bilayers with a convex curvature.