Model building was performed in COOT (Emsley and Cowtan, 2004) based on the x-ray structure of the water-soluble PLY monomer (pdb 5a0d; (van Pee et al., 2016). Initially, individual domains in the x-ray structure of soluble PLY were fitted manually into the cryoEM map as rigid bodies using COOT. Refolded or flexible protein regions were re-fitted manually, followed by geometry regularization in COOT. The complete backbone of PLY was traced in the pore complex map. Densities of bulky side chains were observed in well-ordered regions. The transmembrane β-hairpins, which form upon unfolding of HB1 and HB2 in domain 3 were readily fitted to the map density. The pore model with 42 subunits was generated in UCSF Chimera (Pettersen et al., 2004). Sub-tomogram average maps of the PLY prepore were fitted in UCSF Chimera with the x-ray structures of the PLY monomer (pdb 5cr6; [Marshall et al., 2015]). Sub-tomogram averages of the pore complex were fitted with the cryoEM structure of the membrane-inserted form. Figures were drawn with PyMol (Schrödinger, 2015). FSC curves of model versus map were calculated using the EMAN package (Tang et al., 2007).